Microparticles from Streptococcus pneumoniae as vaccine antigens

ABSTRACT

An isolated Streptococcus pneumoniae membrane vesicle microparticle (MP), wherein said MP comprises: the protein Ply at the level of ≥0.070 μg/μg total protein in the MP; and/or the protein LytA at the level of ≥0.070 μg/μg total protein in the MP; and/or the protein PspC at the level of ≥0.130 μg/μg total protein in the MP; and/or the protein RrgB at the level of ≥0.020 μg/μg total protein in the MP. Compositions comprising such MPs. Uses thereof in particular in immunization, as well as methods of manufacture thereof.

This application is a National Stage application under 35 U.S.C. § 371of International Application No. PCT/SE2017/051323, having anInternational Filing Date of Dec. 21, 2017, which claims the benefit ofSE Application No. 1651746-8 filed Dec. 28, 2016, each of which isincorporated herein by reference in its entirety.

TECHNICAL FIELD

The present invention relates to the field of vaccines againstStreptococcus pneumoniae.

BACKGROUND TO THE INVENTION

Streptococcus pneumoniae is a Gram-positive bacterium that is a majorcontributor to morbidity and mortality worldwide causing about 2 milliondeaths annually, i e the same range as for tuberculosis. Pneumococci arethe major cause of common infections such as sinusitis, otitis, andcommunity-acquired pneumonia (CAP), but also a common cause of severeinvasive diseases (IPD) such as sepsis and meningitis. Young childrenand the elderly are the most susceptible age groups to acquirepneumococcal infections, but other factors such as underlying diseases,splenectomy, immunosuppression (including HIV), diabetes and a priorinfluenza-virus infection also sensitize for a pneumococcal infection.Pneumococcal infections such as CAP also predispose for cardiovasculardiseases such as cardiac infarction, and meningitis patients may getneurological sequelae such as hearing loss and cognitive impairments.

Pneumococci have the human upper airways as their normal ecologicalniche, and up to 60% of preschool children may be colonized at any giventime without symptoms. It is believed that most transmission events tothe elderly come from young colonized children. S. pneumoniae is ahighly genetically diverse species due to an efficient DNA exchangesystem resulting in a multitude of clonal lineages that may differ fromone another in the presence and absence of genes and gene clusters, aswell as in variations in individual genes. Variations in the capsularlocus, the major virulence factor and vaccine target of pneumococci,have resulted in at least 97 known capsular serotypes with differentpotential of causing disease.

Pneumococcal conjugate vaccines (PCVs), targeting a limited number (7,10 or 13 in PCV7, PCV10 and PCV13 respectively) of the known 97 capsularserotypes, have been introduced in the childhood vaccination program inmany countries. Vaccine introduction has led to a decrease in IPD causedby vaccine-types in vaccinated children, however, non-vaccine typepneumococci have rapidly expanded and replaced vaccine-types inchild-hood carriage and in IPD, but also in non-vaccinated populationsuch as the elderly. Furthermore, protection against vaccine typesdiffer, and for serotype 3, a serotype included in PCV13 and with a highmortality rate, there is more or less no evidence for protection againstIPD so far. Data from Sweden, where both PCV10 and PCV 13 are used, showthat serotype 3 was a dominaiting serotype among IPD cases during 2016,also in counties using only PCV13. Hence, novel vaccine approaches areneeded.

In Sweden the total number of patients with IPD has remained almost thesame after vaccine introduction. Moreover, in a recent study we foundthat pneumococcal carriage rates remained around 30% in the youngestchildren 8 years after vaccine introduction in Stockholm, andnon-vaccine types were found in over 90% of the isolates. Antibioticresistance is emerging among clinical pneumococcal isolates andaccording to reports from the European Centre for Disease prevention andControl (ECDC), and the European Antimicrobial Resistance SurveillanceNetwork (EARS-Net), reduced susceptibility to penicillin has increasedduring the last years in Sweden and now exceeds 5% among IPD cases. Inaddition, the need for multiple antigens covering various capsularserotypes lead to high production costs for the known vaccines.

Opsonophagocytosis is the primary mechanism for clearance of pneumococcifrom the host, and the measurement of opsonophagocytic antibodiesappears to correlate with vaccine-induced protection. However, it hasproven difficult to design vaccines that elicit an efficient response inform of opsonophagocytic antibodies.

Taken together, the above issues stress the need for novel vaccineapproaches. Thus, an object of the present invention is the provision ofimproved pneumococcal antigens able to confer serotype-independentimmunological responses and/or improved response in terms ofopsonophagocitic antibodies and protection in experimental models.

Background for Pneumococcal Microparticles

Spherical membranous blebs, so called outer membrane vesicles (OMVs),are produced by Gram-negative bacteria. OMVs range in size from 10 to300 nm and they have been shown to be formed by budding from thebacterial outer membrane, and to have many functions, such asinfluencing virulence by different mechanisms. In Gram-negative bacteriaOMVs have been suggested to act as vehicle to deliver virulence factorsto host cells. Only recently, membrane vesicles (MV_(L)) were discoveredalso in Gram-positive bacteria and their formation and function ispoorly understood compared to OMVs. Gram-positive bacteria do notcontrain an outer membrane and the the cell wall differs fromGram-negative bacteria.

For Streptococcus pneumoniae MV_(L) (membrane vesicle) were isolatedfrom pneumococci belonging to different serotypes (2, 6B, 8 and 23F)grown in liquid culture medium (1). MV_(L) from serotype 8 were shown tobe protective against pneumococcal challenge with the same serotype inmice. Importantly, no cross protection was shown to pneumococci of otherserotypes.

DEFINITIONS

The pneumococcal capsule. The pneumococcal capsule, consisting ofpolysaccharide, exists in at least 97 different variants, so calledcapsular serotypes. The pneumococcal capsule is a major virulencedeterminant of pneumococci, affecting phagocytosis of the bacteria byhost cells, and influence bacterial interactions with the innate immuneresponse.

Pneumolysin (termed Ply herein) is a 53 kDa cholesterol dependentcytolysin released by Streptococcus pneumoniae upon lysis. It is one ofthe major virulence factors of this bacterium. It forms pores in alleukaryotic cells that have cholesterol in their membranes. The formationof pores by Ply frequently results in host cell death as membraneintegrity is destroyed. Ply plays a central role in protecting thepneumococcus from complement attack and aiding its spread to othertissues/organs. Ply is able to activate the classical complementpathway, even in the absence of Ply specific antibody (2). A referencesequence from strain TIGR4 is presented in HQ ID NO: 1.

LytA is the major autolysin of Streptococcus pneumoniae. Lysis is causedby cleaving the lactyl-amide bond between the stem peptides and theglycan strands of peptidoglycan, resulting in hydrolysis of the cellwall. The contribution of LytA to pneumococcal virulence is stillunclear. It is possible that LytA-mediated lysis releases othervirulence factors such as pneumolysin. LytA could also be released tolyse neighboring non-competent pneumococcal cells in a fratricidalmanner. This would potentially facilitate genetic exchange betweennaturally competent pneumococcal populations that easily take up andincorporate DNA by homologous recombination. A third possibility is thatLytA mediates lysis to release proteins involved in immune evasion orcell wall components that may interfere with the host immune response(3). A reference sequence from strain TIGR4 is presented in SEQ ID NO:2.

PspC/CbpA (choline binding protein A) is a protein that binds thephosphocholine present in the teichoic acid and the lipoteichoic acid ofthe cell membrane and the cell wall. It is a major pneumococcal adhesin.It promotes pneumococcal adherence via a human-specific interaction withthe ectodomain of the polymeric Ig receptor. It also prevents activationof C3b and complement-mediated opsonophagocytosis of pneumococci (4).The pspC locus is highly polymorphic and 11 major groups of this proteinhave been identified. Single PspC proteins are identified by sequentialnumbers separated from the group number by a dot (5) as follows:PspC1.1, PspC2.1, PspC2.2, PspC3.1, PspC3.4, PspC4.2/PspC10.1, PspC5.1,PspC6.1/PspC9.1, PspC7.1, PspC8.1, PspC11.1 and PspC11.4 (SEQ ID NOs:3-14, respectively).

RrgB is the major subunit and stalk protein of the pneumococcal pilus.S. pneumoniae pilus 1 is encoded by a genetic islet (PI-1) present in 30to 50% of the pneumococcal strains and is implicated in adhesion toepithelial cells, lung infection, and virulence. Pilus 1 is composed ofthe backbone subunit RrgB, the minor pilin subunits RrgA, and RrgC (6).A reference sequence from strain TIGR4 is presented in SEQ ID NO: 15.

RrgA is a minor pilin subunit of the pneumococcal pilus. RrgA is the tipprotein of the pilus and has been shown to mediate adhesion toepithelial cells (6, 7). Recently, RrgA was shown to promote meningitisdevelopment in mice (8). A reference sequence is presented in SEQ ID NO:16.

PhtD is a polyhistidine triad protein of S. pneumoniae. Polyhistidinetriad (Pht) proteins are streptococcal surface proteins that containmultiple copies of a characteristic HxxHxH sequence, designed ashistidine triads, which were predicted to bind divalent metal cations.Despite the increasing amount of biochemical, structural andphysiological data, the functional role of PhtD and the other Phtproteins remains unclear. PhtD could be involved in zinc uptake.Alternatively, Pht proteins could play a role in protecting thepneumococcus from toxic effects of high Zn²⁺ concentrations byscavenging, storing or trapping Zn²⁺ ions (9). A reference sequence fromstrain TIGR4 is presented in SEQ ID NO: 17.

IgA refers to Immunoglobulin A1 protease of S. pneumoniae (10). SaidIgA1-protease is a highly specific enzyme that cleaves amino acidsequences of certain proteins. The natural substrate of the IgA proteaseis immunoglobulin A1, hence its name. The enzyme is in fact capable ofcleavage of proteins with the amino acid sequenceN—X-Z-Pro-Pro/-Y-Pro-C, where the X in the sequence is preferably aProline or Serine; the Y=Threonine, Serine or Alanine; and Z preferablyis Arginine or Threonine. Thus, the IgA1 protease acts by cleaving theproline-rich hinge region of the heavy chain of IgA1. Release of theIgA1 protease by the pathogen allows adherence to mucous membranes bydegrading host immunoglobulin A. A reference sequence from strain TIGR4is presented in SEQ ID NO: 18.

The term protective immunity in the present context refers toimmunization measures resulting in any degree of reduction in thelikelihood of developing the condition for which the protective immunityis relevant, including a minor, substantial or major reduction inlikelihood of developing the condition as well as total prevention.Preferably, the degree of likelihood reduction is at least a minorreduction.

The term sequence identity expressed in percentage is defined as thevalue determined by comparing two optimally aligned sequences over acomparison window, wherein a portion of the sequence in the comparisonwindow may comprise additions or deletions (i.e., gaps) as compared tothe reference sequence (which does not comprise additions or deletions)for optimal alignment of the two sequences. The percentage is calculatedby determining the number of positions at which the identical amino acidresidue occurs in both sequences to yield the number of matchedpositions, dividing the number of matched positions by the total numberof positions in the window of comparison and multiplying the result by100 to yield the percentage of sequence identity. Unless indicatedotherwise, the comparison window is the entire length of the sequencebeing referred to. In this context, optimal alignment is the alignmentproduced by the BLASTP algorithm as implemented online by the USNational Center for Biotechnology Information (see The NCBI Handbook[Internet], Chapter 16), with the following input parameters: Wordlength=3, Matrix=BLOSUM62, Gap cost=11, Gap extension cost=1.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 : Membrane-derived vesicles and microparticles produced by S.pneumoniae TIGR4 (T4). (A) Electron micrographs and atomic forcemicrographs (insets) of membrane vesicles and microparticles afterisolation and density gradient purification from liquid culture (MV_(L))and microparticles from plate-grown (MP) bacteria. (B) Size distributionof MV_(L), MPs from TIGR4 and MPs from serotype 3. (C) Electronmicrograph of a membrane budding from S. pneumoniae T4R and (D) the sameelectron micrograph in larger magnification. Released particle isindicated by arrow.

FIG. 2 : Identification of proteins present in pneumococcal MV_(L) andMP. (A) Preparations of MV_(L) and MP were separated by SDS-PAGE andproteins visualized by coomassie staining. In comparison, proteinspresent in a whole cell lysate (WC) and supernatant (SN) of a liquidbacterial culture were visualized. (B) Venn diagram displaying thenumber of proteins identified by mass spectrometry (repeated twice)exclusively found in MV_(L) and MP preparations and common to both. (C)Mass spectrometry identification of proteins from MV_(L) and MP. Numbersindicate percentages of proteins based on their subcellularlocalization, defined as cytosolic proteins, membrane-associatedproteins (lipoproteins), proteins with one transmembrane domain(N-terminally anchored), transmembrane proteins, cell wall associatedproteins and secreted proteins. (D) Immunoblot detection of pneumococcalproteins and virulence factors present in MV_(L) and MP isolated from S.pneumoniae T4 WT, pneumolysin-(T4Δply) and LytA-deficient (T4ΔdlytA)strains.

FIG. 3 : Pneumolysin associated with pneumococcal MV_(L) and MP isbiologically active. Quantification of in vitro hemolytic activity ofpurified MV_(L) and MP isolated from S. pneumoniae T4 and T4Δply.Erythrocytes from buffy coat blood were incubated for 60 minutes withdifferent concentrations (0.1, 0.8, 2, 6 and 10 μg protein) of MV_(L)and MP. As control treatments, erythrocytes were incubated with PBS (−)or 1% Triton X-100 (+). Data are represented as means +/−SEM of threeindependent experiments. *=P<0.05; **=P<0.01; ***=P<0.001.

FIG. 4 : Microparticles (MP) are internalized by A549 epithelial cells.

(A, B and C) Immunofluorescence pictures of A549 cells treated with MP(10 μg/ml) for 24 hours. Cells were stained for F-actin, MP are detectedwith anti-pneumolysin and LytA antibodies. (A) Z-stack image (nr.stacks=21) of MP-treated cells. Internalized MP are indicated by arrows.(B and C) Orthogonal views of the same picture, taken at stack nr. 11,for the two indicated MP, further showing their intracellularlocalization.

(D) Immunoblot detection of anti-Ply, and anti-β-actin as loadingcontrol, in A549 cell lysates after incubation with increasingconcentrations (1, 2, 3 μg/ml) of MP. As control treatment, A549 cellswere incubated with PBS (−). (E) Viability of A549 cells examined byflow cytometry of fixable viability dye (FVD) positive cells after 24hours of incubation with MV_(L) or MP. As control treatment, A549 cellswere incubated with PBS (−) or 0.02% NP40 in PBS. Data are representedas means +/−SEM of three independent experiments.

FIG. 5 : MV_(L) and microparticles (MP) activate human monocyte-deriveddendritic cells (DCs). (A) Percentages of FVD and Annexin V-positivecells analyzed by flow cytometry. Cells were incubated for 24 hours withdifferent concentrations (10, 25, 50 μg/ml) of MV₁, MP, PBS (−) or theunencapsulated mutant of TIGR4 (T4R). Data are represented asmeans+/−SEM of three independent experiments. *=P<0.05. (B) Immunoblotdetection of anti-Ply, and anti-3-actin as loading control, in celllysates after incubation of DCs with MP (4 μg/ml) in presence or absenceof the inhibitors Cytochalasin D and Wortmannin (C/W), to block activephagocytosis, or methyl-β-cyclodextrin (MβCD), to block lipid rafts andmembrane fusion. Numbers represent the relative intensity of bandscorresponding to Ply, adjusted to the loading control bands, in threeindependent experiments. (C) DC activation measured by flow cytometry ofMHCII and CD86-positive cells after 24 hours incubation with differentconcentrations (2.5, 5 μg/ml) of MV_(L), MP, PBS (−) or LPS (1 μg/ml).Data are represented as means+/−SEM of three independent experiments.*=P<0.05; ***=P<0.001.

FIG. 6 : Cytokine release by human monocyte-derived dendritic cells uponincubation with MV_(L) or MP. (A) IL-6, (B) IL-8, (C) IL-10 and (D) TNFreleased by dendritic cells after 24 hours of incubation with differentconcentrations (10, 25, 50 μg/ml) of MV_(L) or MP. Control treatmentsinclude PBS (−), the unencapsulated mutant of TIGR4 (T4R) and LPS (1μg/ml). Not detectable (n.d.) amounts of cytokines are indicated. Dataare represented as means+/−SEM of three independent experiments.*=P<0.05; **=P<0.01; ***=P<0.001; ****=P<0.0001.

FIG. 7 : Intranasal immunization of C57BL/6 mice with MV₁ or MP of S.pneumoniae TIGR4 increase survival against intranasal infection withserotype 1.

(A) Percentage of mice survived after challenge. 20 mice per group.*=P<0.05; **=P<0.01; ****=P<0.0001. (B) CFUs in lungs of mice aftersacrifice. Each dot represents one mouse. **=P<0.01; ***=P<0.001. (C-E)CFUs in blood of mice. Each line represents one mouse.

FIG. 8 : Intranasal immunization of mice with MV or MPs stimulate theproduction of pneumoccal-specific IgG. Detection of MW-reactive (A),MP-reactive (B), T4-reactive (C) and T4R-reactive (D) mouse IgG inimmunized mice sera using ELISA assay. Each dot represents one mouseserum. *=P<0.05; **=P<0.01; ***=P<0.001; ****=P<0.0001.

FIG. 9 : Staining of serotypes 1, 3 and 6B using antibodies raisedagainst MP from a serotype 4 strain. (A) Immunofluorescence pictures ofpneumococcal strains belonging to serotypes 1, 3 and 6B stained usingsera, as primary antibodies, from mice immunized either withMV_(L)+adjuvant or MP+adjuvant. The white spots on the bacterial surfaceindicates the presence of specific antibodies against pneumococci in thesera of the immunized mice. (B) Quantification of the signal (whitespots) detected on the bacteria; the signal ratio was calculated bydividing the area of the signal detected on the bacteria (white spots)by the total area detected occupied by the bacteria. The area covered bythe bacteria and the area covered by the signal detected on the bacteriaafter staining with sera were selected, defined and measured using thefunctions Image>Adjust>Threshold and Analyze>Measure of ImageJ.*=P<0.05; **=P<0.01; ***=P<0.001.

FIG. 10 : Opsonophagocytic activity of antibodies in mice sera. (A) RAWmouse macrophages were incubated with Streptococcus pneumoniae type 1,pre-incubated with mouse sera from the adjuvant or MP+adjuvant groups.Indicated are the ratios of adherence of bacteria to cells, or uptake ofbacteria inside cells. Data are represented as means+/−SEM of threeindependent experiments. *=P<0.05. (B) RAW mouse macrophages have beenincubated with Streptococcus pneumoniae type 1, pre-incubated with mousesera from the adjuvant or MP+adjuvant groups. Indicated is thepercentage of taken up bacteria surviving inside the cells after 1 hour.Data are represented as means+/−SEM of three independent experiments.*=P<0.05.

FIG. 11 : Intranasal immunization of C57BL/6 mice with MP of S.pneumoniae increases survival against intranasal infection with serotype3. In particular, immunization with MPs of serotype 3 confers 100%protection against serotype 3 infection in comparison to PCV13immunization, which confers 75% protection. B-cell deficient miceimmunized with MPs of serotype 3 are not protected against intranasalinfection with serotype 3, strongly suggesting that protection providedby MPs is antibody-dependent.

(A) Percentage of mice that survived after pneumococcal challenge. 10mice in the group of “B Cell—(deficient mice)”, 10 mice in the group“Adjuvant”, 20 mice per all other groups. *=P<0.05; ****=P<0.0001. (B)CFUs in lungs of mice after sacrifice. Each dot represents one mouse.***=P<0.001. (C-E) CFUs in blood of mice. Each line represents onemouse.

SUMMARY OF THE INVENTION

The present invention relates to the following items. The subject matterdisclosed in the items below should be regarded disclosed in the samemanner as if the subject matter were disclosed in patent claims.

-   -   1. An isolated Streptococcus pneumoniae membrane vesicle        microparticle (MP), wherein said MP comprises:        -   i. the protein Ply at the level of ≥0.70 μg/μg total protein            in the MP;        -   ii. the protein LytA at the level of ≥0.70 μg/μg total            protein in the MP;        -   iii. the protein PspC at the level of ≥0.130 μg/μg total            protein in the MP; or        -   iv. the protein RrgB at the level of ≥0.020 μg/μg total            protein in the MP.    -   2. The microparticle according to item 1, further comprising a        capsular polysaccharide of a capsular serotype of Streptococcus        pneumoniae, preferably at a level of ≥0.001, more preferably        ≥1.01, most preferably ≥0.1 μg/μg total protein in the MP.    -   3. The microparticle according to any of the preceding items,        comprising the protein Ply at the level of ≥0.70 μg/μg total        protein in the MP.    -   4. The microparticle according to any of the preceding items,        comprising the protein Ply at the level of ≥0.15, preferably        more ≥0.2, preferably most preferably ≥0.35 μg/μg total protein        in the MP.    -   5. The microparticle according to any of the preceding items,        comprising the protein LytA at the level of ≥0.70 μg/μg total        protein in the MP.    -   6. The microparticle according to any of the preceding items,        comprising the protein LytA at the level of ≥0.08, preferably        ≥0.09, yet more preferably ≥0.10, most preferably ≥0.20 μg/μg        total protein in the MP.    -   7. The microparticle according to any of the preceding items,        comprising the protein PspC at the level of ≥0.130 μg/μg total        protein in the MP.    -   8. The microparticle according to any of the preceding items,        comprising the protein PspC at the level of ≥0.15, preferably        ≥0.18, more preferably ≥0.20, most preferably ≥0.3 μg/μg total        protein in the MP.    -   9. The microparticle according to any of the preceding items,        comprising the protein RrgB at the level of ≥0.20 μg/μg total        protein in the MP.    -   10. The microparticle according to any of the preceding items,        comprising the protein RgrB at the level of ≥0.22, preferably        ≥0.025, most preferably ≥0.028 μg/μg total protein in the MP.    -   11. The microparticle according to any of the preceding items,        further comprising the protein PhtD.    -   12. The microparticle according to any of the preceding items,        comprising the protein PhtD at a level being at least 2-fold,        more preferably at least 2.5-fold compared to Streptococcus        pneumoniae membrane vesicles obtained in liquid culture in terms        of μg/μg total protein in the particle or the vesicle,        respectively.    -   13. The microparticle according to any of the preceding items,        further comprising the protein RrgA.    -   14. The microparticle according to any of the preceding items,        comprising the protein RrgA at the level of ≥0.02, preferably        ≥0.05, more preferably ≥0.10, most preferably ≥0.2 μg/μg total        protein in the MP.    -   15. The microparticle according to any of the preceding items,        comprising the protein IgA.    -   16. The microparticle according to any of the preceding items,        comprising the protein IgA at the level of ≥0.02, preferably        ≥0.05, more preferably ≥0.10, most preferably ≥0.2 μg/μg total        protein in the MP.    -   17. The microparticle according to any of the preceding items,        wherein the protein Ply comprises a sequence having at least        80%, preferably 85%, more preferably 90%, yet more preferably        95%, most preferably 100% sequence identity to SEQ ID NO: 1.    -   18. The microparticle according to any of the preceding items,        wherein the protein LytA comprises a sequence having at least        80%, preferably 85%, more preferably 90%, yet more preferably        95%, most preferably 100% sequence identity to SEQ ID NO: 2.    -   19. The microparticle according to any of the preceding items,        wherein the protein PspC comprises a sequence having at least        40%, preferably 80%, more preferably 85%, yet more preferably        90%, still more preferably 95%, most preferably 100% sequence        identity to SEQ ID NOs: 3.    -   20. The microparticle according to any of the preceding items,        wherein the protein PspC comprises a sequence having at least        80%, preferably 85%, more preferably 90%, yet more preferably        95%, most preferably 100% sequence identity to any one of SEQ ID        NOs: 3-14.    -   21. The microparticle according to any of the preceding items,        wherein the protein RgrB comprises a sequence having at least        80%, preferably 85%, more preferably 90%, yet more preferably        95%, most preferably 100% sequence identity to SEQ ID NO: 15.    -   22. The microparticle according to any of the preceding items,        wherein the protein PhtD comprises a sequence having at least        80%, preferably 85%, more preferably 90%, yet more preferably        95%, most preferably 100% sequence identity to SEQ ID NO: 16.    -   23. The microparticle according to any of the preceding items,        comprising the protein RgrA, wherein the protein RgrA comprises        a sequence having at least 80%, preferably 85%, more preferably        90%, yet more preferably 95%, most preferably 100% sequence        identity to SEQ ID NO: 17.    -   24. The microparticle according to any of the preceding items,        comprising the protein IgA, wherein the protein IgA comprises a        sequence having at least 80%, preferably 85%, more preferably        90%, yet more preferably 95%, most preferably 100% sequence        identity to SEQ ID NO: 18.    -   25. The microparticle according to any of the preceding items,        wherein the MP is 5-300 nm in diameter, preferably 15-175 nm.    -   26. The microparticle according to any of the preceding items,        wherein the MP is 10-125 nm in diameter.    -   27. The microparticle according to any of the preceding items,        wherein the MP is derived from a Streptococcus pneumoniae strain        selected from any serotype 1 strain, any serotype 3 strain,        TIGR4, P1031 and A66, preferably TIGR4.    -   28. The microparticle according to any of the preceding items,        obtainable by:        -   a. Culturing host cells of a Streptococcus pneumoniae strain            on blood agar plates;        -   b. harvesting the cultured host cells;        -   c. centrifuging the harvested host cells at 17,000 g for 30            minutes at +4C;        -   d. subjecting the supernatant to filtration through a 0.22            μm filter;        -   e. centrifuging the filtered supernatant at 120,000 g for 2            h at +4C;        -   f. washing the pellets from e) twice in phosphate-buffered            saline with sedimentations at 120,000 g for 2 h at +4C;        -   g. resuspending the pellet in 1 ml phosphate-buffered            saline;        -   h. adjusting the resuspended pellet to 50% (w/v) Optiprep™            density gradient medium in a total volume of 2 ml and            overlaying with one fraction of 30% (w/v) Optiprep™ (9 ml)            followed by a fraction of 5% (w/v) Optiprep™ (3 ml);        -   i. centrifuging the gradients at 250,000×g for 3 hours at 4°            C.;        -   j. collecting the 4 ml top fraction, containing the membrane            microparticle; and        -   k. washing the microparticle three times phosphate-buffered            saline with sedimentations of 250,000×g for 2 hours at +4°            C.    -   29. A composition comprising a microparticle according to any of        items 1-28.    -   30. The composition according to item 29, wherein the        composition is devoid of whole Streptococcus pneumoniae cells.    -   31. The composition according to any of items 29-30, further        comprising capsular polysaccharides from Streptococcus        pneumoniae.    -   32. The composition according to any of items 29-31, comprising        MPs in an amount of 1 μg/ml, preferably 5 μg/ml, more preferably        10 μg/ml, most preferably 100 μg/ml.    -   33. The composition according to any of items 29-32, further        comprising an adjuvant.    -   34. The composition according to any of items 29-33, further        comprising an adjuvant comprising aluminium hydroxide.    -   35. The composition according to any of items 29-34, being an        immunogenic composition.    -   36. The composition according to any of items 29-35, being an        immunogenic composition capable of eliciting opsonophagocitic        antibodies against Streptococcus pneumoniae when administered to        a mammalian host.    -   37. The composition according to any of items 29-36, being an        immunogenic composition is capable of eliciting serotype        independent antibodies against Streptococcus pneumoniae when        administered to a mammalian host.    -   38. The composition according to any of items 29-37, being an        immunogenic composition is capable of eliciting antibodies        against Streptococcus pneumoniae serotype 3 when administered to        a mammalian host.    -   39. The composition according to any of items 29-38, further        comprising capsular polysaccharides of a capsular serotype of        Streptococcus pneumoniae.    -   40. The composition according to any of items 29-39, being a        vaccine.    -   41. The composition according to any of items 29-40, for use in        a method for inducing protective immunity against Streptococcus        pneumoniae in a subject.    -   42. The composition according to item 41, for use according to        item 41, wherein the immunity is protective against a condition        selected from pneumococcal sinusitis, pneumococcal otitis,        pneumococcal pneumonia and invasive pneumococcal disease        including but not limited to pneumococcal sepsis and        pneumococcal meningitis, preferably invasive pneumococcal        disease.    -   43. A use of a microparticle according to any of items 1-28 in        an immunogenic composition.    -   44. The use according to item 43, wherein the immunogenic        composition is a vaccine.    -   45. A use of a microparticle according to any of items 1-28 in        the manufacture of a vaccine.    -   46. A method for manufacturing a vaccine, comprising:        -   a. Providing a microparticle according to any of items 1-28;        -   b. Providing an adjuvant;        -   c. Mixing the microparticle and the adjuvant in a suitable            vehicle in order to produce a vaccine.    -   47. A method for producing an isolated Streptococcus pneumoniae        microparticle according to any of items 1-28, comprising:        -   a. providing a Streptococcus pneumoniae host cell;        -   b. culturing said host cell under conditions allowing the            production of a microparticle according to any of items 1-28            by the host cell; and        -   c. isolating the microparticle thus produced.    -   48. The method according to item 47, wherein the host cell is of        a strain selected from any serotype 1 strain, any serotype 3        strain, TIGR4, P1031 and A66, preferably TIGR4.    -   49. The method according to any of items 47-48, wherein the        culturing is carried out in solid phase.    -   50. The method according to any of items 47-49, wherein the        culturing is carried out in solid phase on blood agar.    -   51. The method according to any of items 47-50, wherein        isolating the microparticle comprises density gradient        centrifugation.

DETAILED DESCRIPTION

The inventors isolated membrane vesicles (MV_(L)) from S. pneumoniae ofserotype 4 grown in liquid cultures as previously described in theliterature (1). They also isolated a novel type of membrane particlestermed microparticles (MP) using a method involving growth of thebacteria overnight on blood agar plates. FIG. 1 shows the appearance ofthe MV_(L) and MP using electron microscopy and atomic force microscopyas well as their size distribution. Microparticles from S. pneumoniaeTIGR4 grown on plates (MP) were smaller in size on average than MV_(L)coming from liquid media (MV_(L)). However, MP from S.pneumoniaeserotype 3 display a different size distribution, containing less of thesmallest particles and more of the largest ones.

Biochemical comparison of MV_(L) and MP preparations from TIGR4 revealeddifferent properties for the MP and MV_(L). MP isolated from platescarried certain pneumococcal proteins to a higher extent than liquidgrown MV_(L). They were particularly enriched in pneumolysin, the majorpore-forming toxin in S. pneumoniae, as shown in FIG. 2D.

The novel MP were characterized using in vitro cell assays, and it wasshown that consistent with the quantitated levels, the MP contain morebiological pneumolysin activity than the MV_(L) (FIG. 3 ). It wasfurther shown that the MP are taken up by epithelial cells (FIG. 4A-D)but are not cytotoxic (FIG. 4E). Using human monocyte-derived dendriticcells it was shown that both MP and MV_(L) are internalized into DCs,induce maturation of DCs (FIG. 5 ), and elicit differential cytokineresponses (FIG. 6 ).

Next, the inventors investigated the potential of MP to protect miceusing experimental models of pneumonia and invasive disease (FIGS. 7-11). The results showed that 80% of mice (wild type C57BL/6 mice)immunized with MP from serotype 4 (combined with aluminium hydroxide asadjuvant) survived after an intranasal pneumococcal infection caused byserotype 1 (heterologous challenge, cross protection). ELISA analysisshowed that mice immunized with MP had a significantly higherMP-specific IgG response in comparison to control mice (treated onlywith the adjuvant) (FIG. 8 ). Immunization with serotype 3 and thenchallenge with serotype 3 led to a 100% protection (FIG. 11 ).Immunization with serotype 4 MP and then challenge with serotype 3 (aserotype that has been problematic in current vaccines) gave aprotection of ca 50% (FIG. 11 ). Furthermore, immunofluorescencestainings showed that sera from mice immunized with MP from serotype 4contained antibodies specific against multiple other serotypes, such asserotypes 1, 3 and 6B (FIG. 9 ).

In conclusion, it was found that MP protect mice against pneumococcalinfection, and, most importantly, such protection isserotype-independent to a significant degree, in contrast to theprotection conferred by currently available vaccines (polysaccharide orconjugated vaccines) that are, as mentioned in the background section,strictly serotype-dependent. The MP immunisation furthermore elicitedprotection against serotype 3 which is a challenge for current vaccines.Indeed, immunization with the currently clinically used vaccine PCV13provided significantly less protection to mice than MP isolated fromserotype 3 (FIG. 11 ).

Moreover, for protein based vaccines that are under development today,there is a problem in obtaining opsonophagocytic capability.Importantly, the present data show that using MP as a vaccine affectsopsonophagocytosis (Example 3, FIG. 10 ). Last but not least, availablepneumococcal vaccines (PCVs) are extremely important to fightpneumococcal diseases in developing countries. However, the costs toproduce current vaccines are very high. An important benefit with thepresent approach is a drop in production costs since isolation ofmicroparticles does not require high-cost preparations withconjugations, instead the method of the present invention is more simplebased on bacterial growth combined with high speed-centrifugation.

Isolated Streptococcus pneumoniae Membrane Vesicles and Microparticles

As discussed in the Background section, a few types of Streptococcuspneumoniae membrane vesicles have been obtained in liquid culture (1).The solid phase culture method used by the inventors resulted in a noveltype of particles termed microparticles having different andadvantageous properties as discussed above and shown in the Examples. Asshown in Table 1, the microparticles differ from the known membranevesicles, MVS in terms of relative protein expression.

TABLE 1 Quantification of proteins present in membrane vesicles, MV_(L),from serotype 4 strain TIGR4 and microparticles, MP, from serotype 4 and3. Percentage sequence identities of protein homologues in referencestrains for serotype 1 (P1031) and serotype 3 (A66) are shown. Theamounts are indicated in μg/μg total protein or relative amount inpercentage. MP MP Homology Homology Protein MV_(L) (TIGR4) (Type 3) inP1031* in A66** Ply 0.055 μg 0.406 μg 0.314 μg 99% 99% LytA 0.050 μg0.100 μg 0.266 μg 99% 100%  PspC 0.118 μg 0.300 μg Not 85% 45% (PspC3.4)determined* (PspC (PspC 2.1) 11.4) RrgB 0.016 μg 0.028 μg Absent AbsentAbsent PhtD 100% 266% 5793% 95% 87% *Because of the high geneticdiversity of the pspC gene among pneumococcal serotypes it was notpossible to use the anti TIGR4 PspC antibody (which was specific for theTIGR4 strain) to detect the serotype 3 PspC

Thus, in a first aspect of the present invention, there is providedisolated Streptococcus pneumoniae microparticle (MP) (a novel type ofpneumococcal membrane vesicle particle), wherein said MP comprise eitherone, two, three or four of the following proteins at the designatedlevels:

-   -   i. the protein Ply at the level of ≥0.070 μg/μg total protein in        the MP;    -   ii. the protein LytA at the level of ≥0.070 μg/μg total protein        in the MP;    -   iii. the protein PspC at the level of ≥0.130 μg/μg total protein        in the MP; and/or    -   iv. the protein RrgB at the level of ≥0.020 μg/μg total protein        in the MP.

The microparticles may further comprise capsular polysaccharides of acapsular serotype of Streptococcus pneumoniae, preferably at a level of≥0.001, more preferably ≥0.01, most preferably ≥0.1 μg/μg total proteinin the MP.

The microparticle may comprise the protein Ply at the level of ≥0.070μg/μg total protein in the MP. The microparticle may comprise theprotein Ply at the level of ≥0.15, preferably ≥0.2, more preferably≥0.3, most preferably ≥0.35 μg/μg total protein in the MP.

The microparticle may comprise the protein LytA at the level of ≥0.070μg/μg total protein in the MP. The microparticle may comprise theprotein LytA at the level of ≥0.08, preferably ≥0.09, yet morepreferably ≥0.1 μg/μg, most preferably ≥0.2 μg/μg total protein in theMP.

The microparticle, MP, may comprise the protein PspC at the level of≥0.130 μg/μg total protein in the MP. The microparticle may comprisecomprising the protein PspC at the level of ≥0.15, preferably ≥0.18,more preferably ≥0.2, most preferably ≥0.3 μg/μg total protein in theMP.

The microparticle may comprise the protein RrgB at the level of ≥0.02μg/μg total protein in the MP. The microparticle may comprise theprotein RgrB at the level of ≥0.022, preferably ≥0.025, most preferably≥0.028 μg/μg total protein in the MP.

The microparticle may further comprise the protein PhtD. The proteinPhtD may be present at a level being at least 2-fold, more preferably atleast 2.5-fold compared to Streptococcus pneumoniae membrane vesiclesobtained in liquid culture in terms of μg/μg total protein in theparticle or the vesicle, respectively.

The microparticle may further comprise the protein RrgA. The proteinRrgA may be present at the level of ≥0.02, preferably ≥0.05, morepreferably ≥0.10, most preferably ≥0.2 μg/μg total protein in the MP.

The microparticle may further comprise the Streptococcus pneumoniaeprotein IgA. The protein IgA may be present at the level of ≥0.02,preferably ≥0.05, more preferably ≥0.10, most preferably ≥0.2 μg/μgtotal protein in the MP.

The protein Ply may comprise a sequence having at least 70%, preferably80%, more preferably 85%, yet more preferably 90%, still more preferably95%, most preferably 100% sequence identity to SEQ ID NO: 1.

The protein LytA may comprise a sequence having at least 70%, preferably80%, more preferably 85%, yet more preferably 90%, still more preferably95%, most preferably 100% sequence identity to SEQ ID NO: 2.

The protein PspC may comprise a sequence having at least 40% or 70%,preferably 80%, more preferably 85%, yet more preferably 90%, still morepreferably 95%, most preferably 100% sequence identity to SEQ ID NOs: 3.

The protein PspC may comprise a sequence having at least 70%, preferably80%, more preferably 85%, yet more preferably 90%, still more preferably95%, most preferably 100% sequence identity to any one of SEQ ID NOs:3-14.

The protein RgrB may comprise a sequence having at least 70%, preferably80%, more preferably 85%, yet more preferably 90%, still more preferably95%, most preferably 100% sequence identity to SEQ ID NO: 15.

The protein PhtD may comprise a sequence having at least 70%, preferably80%, more preferably 85%, yet more preferably 90%, still more preferably95%, most preferably 100% sequence identity to SEQ ID NO: 16.

The protein RgrA may comprise a sequence having at least 70%, preferably80%, more preferably 85%, yet more preferably 90%, still more preferably95%, most preferably 100% sequence identity to SEQ ID NO: 17.

The protein IgA may comprise a sequence having at least 70%, preferably80%, more preferably 85%, yet more preferably 90%, still more preferably95%, most preferably 100% sequence identity to SEQ ID NO: 18.

Thus, the first aspect encompasses (but is not limited to) embodimentsdisclosed in Table 2 below.

TABLE 2 Minimum levels of designated proteins in various embodiments ofthe first aspect. A blank cell indicates that the protein is optionali.e. may be absent or present at any level. Embodiment# Ply LytA PspCPhtD RgrB 1 0.070 2 0.070 0.070 3 0.070 0.130 4 0.070 2-FOLD 5 0.0700.070 0.130 6 0.070 0.070 2-FOLD 7 0.070 0.070 0.130 2-FOLD 8 0.0700.130 2-FOLD 9 0.070 10 0.130 11 2-FOLD 12 0.070 0.130 13 0.070 2-FOLD14 0.070 0.130 2-FOLD 15 0.130 2-FOLD 16 0.070 0.020 17 0.070 0.0700.020 18 0.070 0.130 0.020 19 0.070 2-FOLD 0.020 20 0.070 0.070 0.1300.020 21 0.070 0.070 2-FOLD 0.020 22 0.070 0.070 0.130 2-FOLD 0.020 230.070 0.130 2-FOLD 0.020 24 0.070 0.020 25 0.130 0.020 26 2-FOLD 0.02027 0.070 0.130 0.020 28 0.070 2-FOLD 0.020 29 0.070 0.130 2-FOLD 0.02030 0.130 2-FOLD 0.020 31 0.070 32 0.070 0.070 33 0.070 0.130 34 0.0702-FOLD 35 0.070 0.070 0.130 36 0.070 0.070 2-FOLD 37 0.070 0.070 0.1302-FOLD 38 0.070 0.130 2-FOLD 39 0.070 40 0.130 41 2-FOLD 42 0.070 0.13043 0.070 2-FOLD 44 0.070 0.130 2-FOLD 45 0.130 2-FOLD 46 0.070 0.020 470.070 0.070 0.020 48 0.070 0.130 0.020 49 0.070 2-FOLD 0.020 50 0.0700.070 0.130 0.020 51 0.070 0.070 2-FOLD 0.020 52 0.070 0.070 0.1302-FOLD 0.020 53 0.070 0.130 2-FOLD 0.020 54 0.070 0.020 55 0.130 0.02056 2-FOLD 0.020 57 0.070 0.130 0.020 58 0.070 2-FOLD 0.020 59 0.0700.130 2-FOLD 0.020 60 0.130 2-FOLD 0.020 Units: μg/μg total protein,except for PhtD, where the amount is in comparing to Streptococcuspneumoniae membrane vesicles obtained in liquid culture in terms ofμg/μg total protein in the particle or the vesicle, respectively.

The microparticles may be 5-300 nm in diameter, preferably 10-125 nm indiameter.

The microparticles may be derived from any Streptococcus pneumoniaestrain, but is preferably selected from a group consisting of allserotype 3 strains, all serotype 1 strains, TIGR4, P1031 and A66, mostpreferably TIGR4.

The microparticles according to the first aspect may be obtainable by:

-   -   a. culturing host cells of a Streptococcus pneumoniae strain on        blood agar plates or other plates;    -   b. harvesting the cultured host cells;    -   c. centrifuging the harvested host cells at 17,000 g for 30        minutes at +4° C.;    -   d. subjecting the supernatant to filtration through a 0.22 μm        filter;    -   e. centrifuging the filtered supernatant at 120,000 g for 2 h at        +4° C.;    -   f. washing the pellets from e) twice in phosphate-buffered        saline with sedimentations at 120,000 g for 2 h at +4° C.;    -   g. resuspending the pellet in 1 ml phosphate-buffered saline;    -   h. adjusting the resuspended pellet to 50% (w/v) Optiprep™        density gradient medium in a total volume of 2 ml and overlaying        with one fraction of 30% (w/v) Optiprep™ (9 ml) followed by a        fraction of 5% (w/v) Optiprep™ (3 ml);    -   i. centrifuging the gradients at 250,000×g for 3 hours at 4° C.;    -   j. collecting the 4 ml top fraction, containing the membrane        microparticles;    -   k. washing the microparticles three times phosphate-buffered        saline with sedimentations of 250,000×g for 2 hours at +4° C.

Compositions and Uses

In a second aspect of the present invention, there is provided acomposition comprising a microparticle according to the first aspect.The composition may be devoid of whole Streptococcus pneumoniae cells.The composition may optionally comprise capsular polysaccharides fromStreptococcus pneumoniae.

The composition according to the second aspect may comprise MP in anamount of 1 μg/ml, preferably 5 μg/ml, more preferably 10 μg/ml, mostpreferably 100 μg/ml (the concentrations refer to the total proteincontent of MP per ml).

The composition may further comprise an adjuvant. The adjuvantpreferably comprises aluminium hydroxide.

The composition may be an immunogenic composition. Preferably, theimmunogenic composition is capable of eliciting opsonophagociticantibodies against Streptococcus pneumoniae when administered to amammalian host. Preferably, the immunogenic composition is capable ofeliciting serotype independent antibodies against Streptococcuspneumoniae when administered to a mammalian host. Preferably, theimmunogenic composition is capable of eliciting antibodies againstStreptococcus pneumoniae serotype 3 when administered to a mammalianhost.

The composition may be formulated as a vaccine.

In a third aspect of the present invention, there is provided acomposition according to the second aspect, for use in a method forinducing protective immunity against Streptococcus pneumoniae in asubject. The third aspect also encompasses a method for inducingprotective immunity against Streptococcus pneumoniae in a subject inneed thereof, comprising administering an effective amount of thecomposition according to the second aspect to the subject. The thirdaspect also encompasses the use of a composition according to the secondaspect in the manufacture of a vaccine for immunization againstStreptococcus pneumoniae.

The protective immunity may be an immunity reducing the likelihood of acondition selected from pneumococcal sinusitis, pneumococcal otitis,pneumococcal pneumonia and invasive pneumococcal disease including butnot limited to pneumococcal sepsis and pneumococcal meningitis,preferably invasive pneumococcal disease. Preferably, the subject to beimmunized is a young child (e.g. less than 7 years of age) or an elderlyperson (e.g. over 65 years of age), but also other age groups could betargeted.

For immunization, the composition may be administered to the subject invarious manners known in the art, including but not limited to by way ofinjection (e.g. intramuscular, intracutaneous, subcutaneous,intravenous), buccal, oral and intranasal administration as well asinhalation.

In a fourth aspect there is provided a use of a microparticle accordingto the first aspect in an immunogenic composition. The immunogeniccomposition may be a vaccine.

In a fifth aspect, there is provided a use of a microparticle accordingto the first aspect in the manufacture of a vaccine.

In a sixth aspect, there is provided a method for manufacturing avaccine, comprising:

-   -   a. Providing a microparticle according to the first aspect    -   b. Providing an adjuvant, such as aluminium hydroxide;    -   c. Mixing the microparticle and the adjuvant in a suitable        vehicle in order to produce a vaccine.

Methods for Producing Streptococcus pneumoniae Microparticles

In a sixth aspect of the present invention there is provided a methodfor producing an isolated Streptococcus pneumoniae microparticleaccording to the first aspect, comprising:

-   -   a. providing a Streptococcus pneumoniae bacterial cell;    -   b. culturing said bacterial cell under conditions allowing the        production of a microparticle according to the first aspect by        the bacterial cell; and    -   c. isolating the microparticle thus produced.

The bacterial cell may be from any pneumococcal strain, but ispreferably TIGR4, a serotype 1 and/or a serotype 3 strain.

The culturing may be carried out using plates or liquid culture.Preferably, the culturing is carried out in solid phase, most preferablyon blood agar or other plates. Isolating the microparticles may comprisea density gradient centrifugation step.

General Aspects Relating to the Present Disclosure

The term “comprising” is to be interpreted as including, but not beinglimited to. All references are hereby incorporated by reference. Thearrangement of the present disclosure into sections with headings andsubheadings is merely to improve legibility and is not to be interpretedlimiting in any way, in particular, the division does not in any waypreclude or limit combining features under different headings andsubheadings with each other.

EXAMPLES

The following examples are not to be regarded as limiting. For furtherinformation on the experimental details, the skilled reader is directedto a separate section titled Materials and Methods.

Example 1: Isolation and In Vitro Characterization of Novel PneumococcalMicroparticles

S. pneumoniae TIGR4 Produces Microparticles that Differ Depending onBacterial Growth Conditions.

Membrane vesicles were isolated and purified from Streptococcuspneumoniae serotype 4 strain TIGR4 grown in liquid medium (MV_(L)).Microparticels (MP) were isolated from pneumococci grown on blood agarplates. Preparations were analyzed by transmission electron microscopy(TEM) and atomic force microscopy, and revealed spherical bodiessurrounded by membranous structures (FIG. 1A). The distribution of thesize varied between MV_(L), MP from TIGR4 and MP from serotype 3 (FIG.1B). In total 503 MV_(L), 378 MP from TIGR4 and 523 MP from serotype 3were analyzed. As a control we also studied an isogenic pneumolysinmutant strain (TIGR4Δply) and found a similar size distribution. Toreveal shedding of MV_(L) or MP from the surface of bacteria by TEM wethen used an isogenic unencapsulated mutant in TIGR4 (T4R) grown inliquid culture. In the absence of the thick bacterial capsule, MV_(L)were clearly visible, emanating from the plasma membrane of S.pneumoniae into the extracellular milieu (FIG. 1C-D).

Proteomic Analysis Revealed Differences in Protein Content BetweenMV_(L) from Liquid Culture or MP from Plates

To analyze the protein content of both pneumococcal preparations,purified MV_(L) and MP were subjected to SDS-PAGE analysis. Distinctprotein patterns in MV_(L) and MPs as compared with proteins present inwhole cell lysates or bacterial supernatants (FIG. 2A), were identified.Interestingly, the proteomic content in MV_(L) and MP fractionsdiffered, suggesting a differential enrichment of proteins in thedifferent types of particles. We further conducted proteomic analysisusing tandem mass spectrometry. In total, in TIGR4, we identified 317proteins in MV_(L) (Table 3), and 216 in MP (Table 4), of which 184proteins were found in both preparations. 133 proteins were unique toMV_(L) and 32 to MP (FIG. 2B). In serotype 3 we identified 462 proteinsin MV_(L) (Table 5) and 344 in MP (Table 6). All detected proteins werefurther classified according to their subcellular localization based onthe combination of LocateP and GO annotation (FIG. 2C) (11). A majorityof the proteins identified in MV_(L) and MP were cytosolic proteins,followed by multi-transmembrane proteins, lipoproteins, N-terminallyanchored proteins, secretory proteins and cell wall-associated proteins.While MV_(L) appeared to be more enriched in cytosolic protein and cellwall-associated proteins than MP from TIGR4, the latter harbored ahigher percentage of multi-transmembrane proteins, lipoproteins,N-terminally anchored proteins and secretory proteins.

We further analyzed the mass spectrometry data in order to determine thepresence of 30 known pneumococcal virulence factors in the particles.Both types of particles contained approximately half of these virulencefactors present in TIGR4. Just two were unique to MV_(L), CbpD (12) andBgaA (13), and one to MP, IgA (10).

Based on the proteomic results, we next performed immunoblot analysis onspecific pneumococcal proteins and virulence factors (FIG. 2D). Amongcytosolic proteins we investigated the presence of the cytotoxinpneumolysin (Ply), which is a pore-forming toxin and one of the mostimportant virulence factors in pneumococcal pathogenesis (2), GAPDH as acytosolic marker (14), and LytA, the major autolysin of Streptococcuspneumoniae (3).

The most striking observation was the high enrichment of Ply especiallyin MP, which also contained more LytA than MV_(L). Smaller differenceswere also observed in the enrichment of virulence factors belonging toother subcellular localizations. While MV_(L) displayed a higher amountof Pneumococcal surface antigen A (PsaA) (15) and Sortase A (SrtA) (16)than MP, the latter contained more Pneumococcal surface protein C (PspC)(17) and Polyhistidine triad protein D (PhtD) (9) (FIG. 2D). The majorcomponent of the pneumococcal pilus, RrgB (6), was equally found in bothparticle preparations.

These findings suggest the presence of a still uncharacterized mechanismfor the selective secretion and enrichment of specific proteins andvirulence factors of Streptococcus pneumoniae in these particlepreparations.

A Biologically Active Pneumolysin was Found in Membrane Microparticles

The enrichment of Ply in both MV_(L) and MP prompted us to determinewhether particle-associated Ply was functional and able to lyseerythrocytes in a hemolytic assay. For both types of particles weobserved a dose dependent hemolysis (FIG. 3 ), however, Ply from MPshowed higher activity than from MV_(L), which is consistent with theobservation that MP contain more Ply. Notably, hemolysis seemed to beentirely caused by Ply, and particles prepared from a TIGR4 mutantlacking Ply (TIGR4Δply) exhibited no hemolytic activity.

MP are Taken Up by A549 Epithelial Cells, but are not Cytotoxic.

MV in other Gram-positive bacteria have been described to function asvehicle to deliver vesicle-associated proteins to human cells (18) (19).To test whether pneumococcal MP can deliver pneumococcal proteins, weincubated MP from TIGR4 with A549 lung epithelial cells for 24 hours andvisualized Ply and LytA by immunofluorescence staining (FIG. 4A-C). Byanalyzing orthogonal views of A549 cells treated with MP, we showed byimmunofluorescence that MP can be taken up, with Ply and LytA detectableinside the cells (FIG. 4B-C).

To confirm our observation we further analyzed lysates of A549 cellstreated with MP by immunoblotting with anti-Ply antibody (FIG. 4D). Inorder to minimize extracellular associated particles, treated cells wereexcessively washed before lysates were made. Consistent with ourimmunofluorescence results, western blotting revealed a dose dependentincrease in Ply further suggesting that MP can be internalized.

Next we asked whether proteins associated with MV_(L) or MP, and inparticular Ply, may contribute to cytotoxic effects on cultured humancells. To assess toxicity, A549 cells were treated with differentconcentrations of MV_(L) and MP for 24 hours, stained with fixableviability dye and analyzed by flow cytometry (FIG. 4E). The differencein cell death between samples incubated with MV_(L) or MPs and thenegative control sample was not statistically significant, suggestingthat Ply is not cytotoxic to the cells at concentrations present inMV_(L) or MP.

MV_(L) and MP are Internalized by Human Monocyte-Derived Dendritic Cells(DCs) and Induce their Maturation.

Since we found that MP can be internalized by epithelial cells, we thenstudied potential interactions of MV_(L) or MP with innate immune cells,focusing on DCs. First we assessed cytotoxicity by incubating DCs withMV_(L) or MP, stained with fixable viability dye and Annexin V, and thenanalyzed using flow cytometry (FIG. 5A). No significant increase incytotoxicity was detected in samples incubated with MP, while additionof MV_(L) displayed cytotoxic effects at the two highest concentrationsused. While both preparations were considerably less cytotoxic than livebacteria, with 70-80% cytotoxic cells after 24 hours of incubation, morestudies are needed to assess the mechanism behind the increasedcytotoxicity of MV_(L) from liquid cultures. Moreover, the percentage ofapoptotic cells did not change after incubation with either of theparticle preparations.

Then we investigated uptake of MP by DCs. First we incubated DCs with MPfor 1 hour, lysed the cells and performed SDS-PAGE and immunoblotting oncell lysates to check for presence of Ply. Indeed, Ply was found in thecell lysates (FIG. 5B), suggesting that DCs internalize MP. Next, DCswere challenged for 30 minutes with inhibitors of actin polymerizationand phosphatidylinositol 3-kinase (PI3K), cytochalasin and wortmannin,to block active phagocytosis, or with an inhibitor of lipid rafts,methyl-β-cyclodextrin, to block fusion of membranes. Subsequently, cellswere incubated with MP for 1 hour, lysed, and the same protocol forimmunoblotting was followed. Interestingly, blocking active phagocytosisor membrane fusion resulted in less Ply. However, when all inhibitorswere used together, Ply still remained present in the lysates,suggesting either that the particles adhered to the cells or wereinternalized by another mechanism, not investigated in this study.

Taken together these data suggest that MP can function as a vehicle todeliver bacterial components to host cells. Since DCs are professionalantigen presenting cells, we then assessed whether MV_(L) or MP couldinfluence DC maturation. Cells were incubated for 24 hours with MV_(L)or MP, and then stained for MHCII and the co-stimulatory molecule CD86,and analyzed by flow cytometry (FIG. 5C). A dose-dependent increase inthe presence of both markers on the surface of DCs was detected whencells were incubated with the particles. MV_(L) appeared to induce morematuration than MP, and similar levels of maturation were obtained withLPS.

MV_(L) and MP Induce Pro-Inflammatory Cytokine Responses in DCs

Since OMVs from Gram-negative have been shown to trigger a potent innateimmune response (20) (21), we studied whether also pneumococcal MV_(L)or MP could affect pro-inflammatory cytokine responses. DCs we incubatedwith MV_(L) or MP for 24 hours, and ELISA assays were used onsupernatants to determine concentrations of IL-6 (FIG. 6A), IL-8 (FIG.6B), IL-10 (FIG. 6C) and TNF (FIG. 6D). MP induced release of IL-8 andlow levels of TNF, while no induction was seen for IL-6 and IL-10. Incontrast, MV_(L) induced all four cytokines tested to levels higher thanlive bacteria. None of the particles induced IL-10 or IL-12 (data notshown).

TABLE 3 Proteins identified in MV_(L) from TIGR4. The accession numbersrefer to SwissProt on the date of filing. Accession Coverage # Peptides#AAs Score Desription Localization Q97RH0 76.86 47 350 2615.53Lipoprotein OS = Streptococcus Lipid pneumoniae serotype 4 (strain ATCCanchored BAA-334/TIGR4) GN = SP_0845 PE = 4 SV = 1 − [Q97RH0_STRPN]Q2MGF6 82.45 59 490 1620.74 Lysozyme OS = Streptococcus Secretorypneumoniae serotype 4 (strain ATCC (released) BAA-334/TIGR4) GN = lytCPE = 4 SV = 1 − (with CS) [Q2MGF6_STRPN] Q97NK0 73.39 64 883 1364.43Aldehyde-alcohol dehydrogenase Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_2026 PE = 3SV = 1 − [Q97NK0_STRPN] P18791 72.99 48 659 1307.17 Oligopeptide-bindingprotein AmiA Lipid OS = Streptococcus pneumoniae anchored serotype 4(strain ATCC BAA-334/ TIGR4) GN = amiA PE = 1 SV = 3 − [AMIA_STRPN]Q9L7Q2 55.82 91 1906 1166.89 Zinc metalloprotease ZmpB Multi- OS =Streptococcus pneumoniae trans- serotype 4 (strain ATCC BAA-334/membrane TIGR4) GN = zmpB PE = 3 SV = 2 − [ZMPB_STRPN] Q97T80 58.3 871856 1012.57 Zinc metalloprotease ZmpC LPxTG OS = Streptococcuspneumoniae Cell-wall serotype 4 (strain ATCC BAA-334/ anchored TIGR4) GN= zmpC PE = 3 SV = 1 − [ZMPC_STRPN] Q97NQ8 59.59 75 1225 936.07DNA-directed RNA polymerase subunit Intracellular beta′ OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =rpoC PE = 3 SV = 1 − [RPOC_STRPN] P59213 74.47 48 423 838.2Maltose/maltodextrin-binding protein Lipid OS = Streptococcus pneumoniaeanchored serotype 4 (strain ATCC BAA-334/ TIGR4) GN = malX PE = 1 SV = 1− [MALX_STRPN] P64022 75.61 46 693 803.77 Elongation factor G OS =Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = fusA PE = 3 SV = 1 − [EFG_STRPN] P0C2J9 92.99 46 471753.38 Pneumolysin OS = Streptococcus Intracellular pneumoniae serotype4 (strain ATCC BAA-334/TIGR4) GN = ply PE = 3 SV = 1 − [TACY_STRPN]Q97N55 75 27 392 729.12 Secreted 45 kd protein Secretory OS =Streptococcus pneumoniae (released) serotype 4 (strain ATCC BAA-334/(with CS) TIGR4) GN = usp45 PE = 4 SV = 1 − [Q97N55_STRPN] Q97RY6 42.173 2140 715.85 Serine protease, subtilase family LPxTG OS =Streptococcus pneumoniae Cell-wall serotype 4 (strain ATCC BAA-334/anchored TIGR4) GN = SP_0641 PE = 4 SV = 1 − [Q97RY6_STRPN] P64030 68.8432 398 684.61 Elongation factor Tu Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = tuf PE = 3 SV =1 − [EFTU_STRPN] Q97NQ7 66.42 61 1203 683.01 DNA-directed RNA polymerasesubunit Intracellular beta OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = rpoB PE = 3 SV = 1 − [RPOB_STRPN]Q97PE6 77.38 35 442 645.41 Sugar ABC transporter, sugar-binding Lipidprotein OS = Streptococcus anchored pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_1683 PE = 4 SV = 1 − [Q97PE6_STRPN] Q97PT1 74.5945 551 641.52 Uncharacterized protein Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_1518 PE = 4SV = 1 − [Q97PT1_STRPN] Q97RQ0 81.35 26 386 599.79 Branched-chain aminoacid ABC Lipid transporter, amino acid-binding anchored protein OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =livJ PE = 4 SV = 1 − [Q97RQ0_STRPN] Q97SV2 62.45 19 277 575.15 50Sribosomal protein L2 Intracellular OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = rplB PE = 3 SV = 1 −[RL2_STRPN] Q97R51 74.12 32 313 574.61 Foldase protein PrsA Lipid OS =Streptococcus pneumoniae anchored serotype 4 (strain ATCC BAA-334/TIGR4) GN = prsA PE = 3 SV = 1 − [PRSA_STRPN] Q97SV5 59.62 17 208 571.9250S ribosomal protein L3 Intracellular OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = rplC PE = 3 SV = 1 −[RL3_STRPN] Q04707 64.12 38 719 564.47 Penicillin-binding protein 1A N-OS = Streptococcus pneumoniae terminally serotype 4 (strain ATCCBAA-334/ anchored TIGR4) GN = ponA PE = 1 SV = 2 − (No CS) [PBPA_STRPN]Q97NL1 88.36 26 335 546.25 Glyceraldehyde-3-phosphate Intracellulardehydrogenase OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = gap PE = 3 SV = 1 − [Q97NU_STRPN] Q97PD6 64.87 46837 501.57 Protein translocase subunit SecA 1 Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =secA1 PE = 3 SV = 1 − [SECA1_STRPN] P35592 75.61 44 660 500.22Oligopeptide-binding protein AliA Lipid OS = Streptococcus pneumoniaeanchored serotype 4 (strain ATCC BAA-334/ TIGR4) GN = aliA PE = 3 SV = 4− [ALIA_STRPN] P0A4G2 72.49 21 309 482.72 Manganese ABC transporterLipid substrate-binding lipoprotein anchored OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = psaA PE = 1 SV =1 − [MTSA_STRPN] Q54970 61.59 38 591 482.11 Pyruvate oxidase OS =Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = spxB PE = 3 SV = 2 − [PDXB_STRPN] Q97R09 68.62 21341 442.02 Iron-compound ABC transporter, iron Lipid compound-bindingprotein anchored OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = SP_1032 PE = 1 SV = 1 − [Q97R09_STRPN] Q97PU3 67.6331 278 434.28 Amino acid ABC transporter, amino Lipid acid-bindingprotein anchored OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = aatB PE = 4 SV = 1 − [Q97PU3_STRPN] P0A4C3 72.35 20217 427.97 30S ribosomal protein S3 Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = rpsC PE = 3 SV =1 − [RS3_STRPN] Q97T39 36.42 28 744 407.46 Pneumococcal surface proteinA N- OS = Streptococcus pneumoniae terminally serotype 4 (strain ATCCBAA-334/ anchored TIGR4) GN = pspA PE = 4 SV = 1 − (with CS)[Q97T39_STRPN] I6L8V7 41.79 28 627 381.01 Choline binding protein ESecretory OS = Streptococcus pneumoniae (released) serotype 4 (strainATCC BAA-334/ (with CS) TIGR4) GN = cbpE PE = 4 SV = 1 − [I6L8V7_STRPN]P0A475 54.74 11 137 354.71 50S ribosomal protein L16 Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =rplP PE = 1 SV = 1 − [RL16_STRPN] P66112 51.26 9 119 346.49 50Sribosomal protein L20 Intracellular OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = rplT PE = 3 SV = 1 −[RL20_STRPN] Q97SV4 56.04 14 207 345.01 50S ribosomal protein L4Intracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = rplD PE = 3 SV = 1 − [RL4_STRPN] Q97T12 78.99 26276 332.96 ABC transporter, substrate-binding Lipid protein OS =Streptococcus anchored pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4)GN = SP_0148 PE = 4 SV = 1 − [Q97T12_STRPN] Q97N74 47.47 31 693 320.12Choline binding protein A N- OS = Streptococcus pneumoniae terminallyserotype 4 (strain ATCC BAA-334/ anchored TIGR4) GN = cbpA PE = 1 SV = 1− (with CS) [Q97N74_STRPN] Q97QS2 58.06 22 434 315.73 Enolase OS =Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = eno PE = 1 SV = 1 − [ENO_STRPN] Q97S57 49.58 34 958315.37 Translation initiation factor IF-2 Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =infB PE = 3 SV = 1 − [IF2_STRPN] Q97T11 75 27 284 313.6 Lipoprotein OS =Streptococcus Lipid pneumoniae serotype 4 (strain ATCC anchoredBAA-334/TIGR4) GN = SP_0149 PE = 3 SV = 1 − [Q97T11_STRPN] P66359 55.1210 127 299.97 30S ribosomal protein S11 Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = rpsK PE = 3 SV =1 − [RS11_STRPN] Q97PT6 68.8 24 468 285.18 ATP synthase subunit betaIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = atpD PE = 3 SV = 1 − [ATPB_STRPN] Q97SP2 60.24 18332 279.63 PTS system, mannose-specific IIAB Intracellular components OS= Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =manL PE = 4 SV = 1 − [Q97SP2_STRPN] Q97RZ7 60.08 15 238 278.06Uncharacterized protein Lipid OS = Streptococcus pneumoniae anchoredserotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_0629 PE = 1 SV = 1 −[Q97RZ7_STRPN] Q97NL3 55.13 34 731 275.05 Penicillin-binding protein 2AN- OS = Streptococcus pneumoniae terminally serotype 4 (strain ATCCBAA-334/ anchored TIGR4) GN = pbp2A PE = 4 SV = 1 − (No CS)[Q97NL3_STRPN] Q97PE1 62.92 29 445 269.34 ABC transporter,substrate-binding Lipid protein OS = Streptococcus anchored pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1690 PE = 4 SV = 1 −[Q97PE1_STRPN] Q97SX2 81.48 19 189 268.65 Uncharacterized protein LipidOS = Streptococcus pneumoniae anchored serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_0191 PE = 1 SV = 1 − [Q97SX2_STRPN] Q97RN2 67.79 15 267248.33 Peptidyl-prolyl cis-trans isomerase, Lipid cyclophilin-type OS =Streptococcus anchored pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4)GN = SP_0771 PE = 3 SV = 1 − [Q97RN2_STRPN] P65887 62.62 25 428 243.93Adenylosuccinate synthetase Intracellular OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = purA PE = 1 SV = 1 −[PURA_STRPN] Q97QH2 33.7 23 721 242.51 Amino acid ABC transporter, aminoMulti- acid-binding protein/permease trans- protein OS = Streptococcusmembrane pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1241PE = 1 SV = 1 − [Q97QH2_STRPN] O69076 50.46 25 652 240.51 ATP-dependentzinc metalloprotease Multi- FtsH OS = Streptococcus pneumoniae trans-serotype 4 (strain ATCC BAA-334/ membrane TIGR4) GN = ftsH PE = 3 SV = 3− [FTSH_STRPN] Q97N37 62.6 22 393 236.55 Serine protease OS =Streptococcus N- pneumoniae serotype 4 (strain ATCC terminallyBAA-334/TIGR4) GN = SP_2239 PE = 1 anchored SV = 1 − [Q97N37_STRPN] (NoCS) Q97T63 60.29 26 491 232.33 ABC transporter, substrate-binding Lipidprotein OS = Streptococcus anchored pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_0092 PE = 4 SV = 1 − [Q97T63_STRPN] Q97NQ0 68.7919 330 230.07 Aspartate-ammonia ligase Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = asnA PE = 3 SV =1 − [ASNA_STRPN] Q97SV1 86.67 12 180 229.35 50S ribosomal protein L5Intracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = rplE PE = 3 SV = 1 − [RL5_STRPN] Q97Q37 69.37 18271 227.84 Amino acid ABC transporter, amino Lipid acid-binding proteinanchored OS = Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1394 PE = 4 SV = 1 − [Q97Q37_STRPN] P66907 42.63 9 380223.83 Queuine tRNA-ribosyltransferase Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = tgt PE = 3 SV =1 − [TGT_STRPN] P59205 43.77 20 658 217.64 Putative endo-beta-N-Secretory acetylglucosaminidase (released) OS = Streptococcus pneumoniae(with CS) serotype 4 (strain ATCC BAA-334/ TIGR4) GN = lytB PE = 1 SV =1 − [LYTB_STRPN] Q97SC6 49.35 27 774 216.35 Formate acetyltransferaseIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = pfl PE = 4 SV = 1 − [Q97SC6_STRPN] Q97NB5 42.19 22621 215.44 Choline binding protein PcpA Secretory OS = Streptococcuspneumoniae (released) serotype 4 (strain ATCC BAA-334/ (with CS) TIGR4)GN = pcpA PE = 1 SV = 1 − [Q97NB5_STRPN] P0A2U8 49.58 19 355 215.01Oligopeptide transport ATP-binding Intracellular protein AmiE OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =amiE PE = 3 SV = 1 − [AMIE_STRPN] Q97PA9 39 21 659 212.01Serine/threonine-protein kinase StkP Intracellular/ OS = Streptococcuspneumoniae TMH serotype 4 (strain ATCC BAA-334/ start TIGR4) GN = stkPPE = 1 SV = 1 − AFTER 60 [STKP2_STRPN] Q97RF9 56.25 22 400 204.01Ribosomal protein S1 Intracellular OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = rpsA PE = 4 SV = 1 −[Q97RF9_STRPN] Q97SP4 43.89 12 303 202.88 PTS system, mannose-specificIID Multi- component OS = Streptococcus trans- pneumoniae serotype 4(strain ATCC membrane BAA-334/TIGR4) GN = SP_0282 PE = 4 SV = 1 −[Q97SP4_STRPN] P0A4A7 58.39 11 137 202.86 30S ribosomal protein S12Intracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = rpsL PE = 3 SV = 1 − [RS12_STRPN] Q97Q31 55.82 13292 202.43 Phosphate-binding protein PstS 1 Lipid OS = Streptococcuspneumoniae anchored serotype 4 (strain ATCC BAA-334/ TIGR4) GN = pstS1PE = 1 SV = 1 − [PSTS1_STRPN] Q97N56 69.11 20 259 201.44 30S ribosomalprotein S2 Intracellular OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = rpsB PE = 3 SV = 1 − [RS2_STRPN]Q97RC6 48.66 14 335 201.21 ATP-dependent 6- Intracellularphosphofructokinase OS = Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/ TIGR4) GN = pfkA PE = 3 SV = 1 − [PFKA_STRPN] Q97QX5 68.0215 344 200.45 Uncharacterized protein Secretory OS = Streptococcuspneumoniae (released) serotype 4 (strain ATCC BAA-334/ (with CS) TIGR4)GN = SP_1069 PE = 1 SV = 1 − [Q97QX5_STRPN] Q97NM6 60.64 17 404 197.5Aminotransferase, class I Intracellular OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_1994 PE = 4 SV = 1 −[Q97NM6_STRPN] Q97N69 65.37 19 335 194.36 Putative ABC transporter,substrate- Lipid binding protein OS = Streptococcus anchored pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_2197 PE = 4 SV = 1 −[Q97N69_STRPN] Q97NS5 60.95 16 338 193.02 Putative transcriptionalregulator Secretory OS = Streptococcus pneumoniae (released) serotype 4(strain ATCC BAA-334/ (with CS) TIGR4) GN = SP_1942 PE = 4 SV = 1 −[Q97NS5_STRPN] P0A4M7 38.35 20 498 189.98 Oligopeptide transport systemMulti- permease protein AmiC transmem- OS = Streptococcus pneumoniaebrane serotype 4 (strain ATCC BAA-334/ TIGR4) GN = amiC PE = 3 SV = 1 −[AMIC_STRPN] Q97QD2 56.41 20 523 188.15 Signal recognition particleprotein Intracellular OS = Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/ TIGR4) GN = ffh PE = 1 SV = 1 − [Q97QD2_STRPN] P1467749.87 26 750 185.33 Penicillin-binding protein 2 × N- OS = Streptococcuspneumoniae terminally serotype 4 (strain ATCC BAA-334/ anchored TIGR4)GN = pbpX PE = 1 SV = 2 − (No CS) [PBPX_STRPN] Q97NQ3 51.88 14 345183.26 Uncharacterized protein N- OS = Streptococcus pneumoniaeterminally serotype 4 (strain ATCC BAA-334/ anchored TIGR4) GN = SP_1967PE = 4 SV = 1 − (No CS) [Q97NQ3_STRPN] Q97RW9 59.34 16 332 182.68Putative pneumococcal surface Lipid protein OS = Streptococcus anchoredpneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_0667 PE = 1 SV= 1 − [Q97RW9_STRPN] Q97RL9 66.17 16 399 180.86 Uncharacterized proteinN- OS = Streptococcus pneumoniae terminally serotype 4 (strain ATCCBAA-334/ anchored TIGR4) GN = SP_0785 PE = 4 SV = 1 − (No CS)[Q97RL9_STRPN] P0A3M9 47.26 14 328 180.34 L-lactate dehydrogenaseIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = ldh PE = 3 SV = 2 − [LDH_STRPN] Q97PT4 39.72 17 501180.17 ATP synthase subunit alpha Multi- OS = Streptococcus pneumoniaetrans- serotype 4 (strain ATCC BAA-334/ membrane TIGR4) GN = atpA PE = 3SV = 1 − [ATPA_STRPN] Q97QH1 67.48 9 246 175.25 Amino acid ABCtransporter, ATP- Intracellular binding protein OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1242 PE = 3 SV= 1 − [Q97QH1_STRPN] Q97NX9 72.9 19 321 174.95 Iron-compound ABCtransporter, iron- Lipid compound-binding protein anchored OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =SP_1872 PE = 4 SV = 1 − [Q97NX9_STRPN] Q97QC6 66.09 14 115 173.94 50Sribosomal protein L19 Intracellular OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = rplS PE = 3 SV = 1 −[RL19_STRPN] Q97N99 63.14 18 274 172.57 SPFH domain/Band 7 familyIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = SP_2156 PE = 4 SV = 1 − [Q97N99_STRPN] Q97PW1 50.9717 463 169.74 Peptidoglycan N-acetylglucosamine N- deacetylase A OS =Streptococcus terminally pneumoniae serotype 4 (strain ATCC anchoredBAA-334/TIGR4) GN = pgdA PE = 4 SV = 1 − (No CS) [Q97PW1_STRPN] Q97SE136.4 14 555 169.45 Uncharacterized protein Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =SP_0443 PE = 1 SV = 1 − [Q97SE1_STRPN] Q97PM5 59.04 16 376 168.92 SugarABC transporter, ATP-binding Intracellular protein OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = msmK PE = 4 SV =1 − [Q97PM5_STRPN] Q97SN5 85.81 15 148 165.86 50S ribosomal protein L13Intracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = rplM PE = 3 SV = 1 − [Q97SN5_STRPN] Q97PQ2 28.88 18914 162.85 Cation-transporting ATPase, E1-E2 Multi- family OS =Streptococcus pneumoniae trans- serotype 4 (strain ATCC BAA-334/membrane TIGR4) GN = SP_1551 PE = 3 SV = 1 − [Q97PQ2_STRPN] Q97NE4 32.1621 821 161.61 Penicillin-binding protein 1B N- OS = Streptococcuspneumoniae terminally serotype 4 (strain ATCC BAA-334/ anchored TIGR4)GN = pbp1B PE = 4 SV = 1 − (No CS) [Q97NE4_STRPN] Q97Q62 54.48 20 424161.35 Psr protein OS = Streptococcus Intracellular/ pneumoniae serotype4 (strain ATCC TMH BAA-334/TIGR4) GN = SP_1368 PE = 4 start SV = 1 −[Q97Q62_STRPN] AFTER 60 P18766 50 13 308 160.11 Oligopeptide transportATP-binding Intracellular protein AmiF OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = amiF PE = 3 SV = 2 −[AMIF_STRPN] I6L8U0 42.39 16 394 155.34 UDP-N-acetylglucosamine 2-Intracellular epimerase OS = Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/TIGR4) GN = cps4L PE = 3 SV = 1 − [I6L8U0_STRPN] P0CC0855.21 15 288 154.74 Acetyl-coenzyme A carboxylase Intracellular carboxyltransferase subunit beta OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = accD PE = 1 SV = 1 − [ACCD_STRPN]Q97RG9 51.27 21 511 154.04 Sugar ABC transporter, ATP-bindingIntracellular protein OS = Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/TIGR4) GN = SP_0846 PE = 3 SV = 1 − [Q97RG9_STRPN] Q97SJ450.28 17 360 153.5 Uncharacterized protein Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =SP_0355 PE = 4 SV = 1 − [Q975J4_STRPN] P72524 33.82 22 822 148.92 DNAgyrase subunit A Intracellular OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = gyrA PE = 3 SV = 3 − [GYRA_STRPN]P66565 56.16 17 203 148.91 30S ribosomal protein S4 Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =rpsD PE = 3 SV = 1 − [RS4_STRPN] P0A4G0 41.1 20 652 146.87Oligopeptide-binding protein AliB Lipid OS = Streptococcus pneumoniaeanchored serotype 4 (strain ATCC BAA-334/ TIGR4) GN = aliB PE = 3 SV = 1− [ALIB_STRPN] Q97SG0 57.78 18 424 144.79 Serine-tRNA ligase OS =Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = serS PE = 3 SV = 1 − [SYS_STRPN] Q97PF9 51.79 20 419144.46 Cell division protein FtsZ Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = ftsZ PE = 3 SV =1 − [Q97PF9_STRPN] Q97Q34 64.04 17 267 144.16 Phosphate importATP-binding protein Intracellular PstB 2 OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = pstB2 PE = 3 SV = 1 −[PSTB2_STRPN] I6L8N1 47.25 14 455 141.81 Acetyl-CoA carboxylase, biotinIntracellular carboxylase OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = accC PE = 4 SV = 1 − [I6L8N1_STRPN]Q97SR1 45.54 19 617 140.84 Proline-tRNA ligase OS = StreptococcusIntracellular pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =proS PE = 3 SV = 1 − [SYP_STRPN] Q97SI4 40.88 11 340 140.1 Cholinebinding protein C N- OS = Streptococcus pneumoniae terminally serotype 4(strain ATCC BAA-334/ anchored TIGR4) GN = cbpC PE = 4 SV = 1 − (withCS) [Q97SI4_STRPN] P0A4D7 40.84 17 524 138.63 DEAD-box ATP-dependent RNAIntracellular helicase CshA OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = cshA PE = 3 SV = 1 − [CSHA_STRPN]Q97RN3 44.44 17 513 136.66 ABC transporter, ATP-binding proteinIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = SP_0770 PE = 1 SV = 1 − [Q97RN3_STRPN] Q97PR0 43.6217 447 136.28 Asparagine-tRNA ligase Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = asnS PE = 1 SV =1 − [SYN_STRPN] P22976 40.73 18 658 132.04 Probable transketolaseIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = tkt PE = 3 SV = 2 − [TKT_STRPN] Q97PX1 34.2 16 459132 NADH oxidase OS = Streptococcus Intracellular pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = nox PE = 4 SV = 1 − [Q97PX1_STRPN]O05703 50.5 17 501 129.61 Zinc-binding lipoprotein AdcA Lipid OS =Streptococcus pneumoniae anchored serotype 4 (strain ATCC BAA-334/TIGR4) GN = adcA PE = 3 SV = 4 − [ADCA_STRPN] P66095 52.4 12 229 128.4450S ribosomal protein L1 Intracellular OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = rplA PE = 3 SV = 1 −[RL1_STRPN] Q97RP4 55.65 12 230 127.75 Cell division ABC transporter,ATP- Intracellular binding protein FtsE OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = ftsE PE = 3 SV = 1 −[Q97RP4_STRPN] Q97RP3 37.66 12 308 126.99 Cell division protein FtsXMulti- OS = Streptococcus pneumoniae trans- serotype 4 (strain ATCCBAA-334/ membrane TIGR4) GN = ftsX PE = 3 SV = 1 − [Q97RP3_STRPN] Q97N7238.4 23 810 122.4 ATP-dependent Clp protease, ATP- Intracellular bindingsubunit OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_2194 PE = 3 SV = 1 − [Q97N72_STRPN] Q97Q67 43.914 344 121.11 Conserved domain protein Multi- OS = Streptococcuspneumoniae trans- serotype 4 (strain ATCC BAA-334/ membrane TIGR4) GN =SP_1363 PE = 4 SV = 1 − [Q97Q67_STRPN] P95830 50.79 15 378 120.38Chaperone protein DnaJ Intracellular OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = dnaJ PE = 1 SV = 2 −[DNAJ_STRPN] Q97SJ6 53.04 8 230 119.51 Capsular polysaccharidebiosynthesis Multi- protein CpsC OS = Streptococcus trans- pneumoniaeserotype 4 (strain ATCC membrane BAA-334/TIGR4) GN = cpsC PE = 3 SV = 1− [CPSC_STRPN] P63373 71.03 13 252 116.79 Phosphate import ATP-bindingprotein Intracellular PstB 1 OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = pstB1 PE = 3 SV = 1 − [PSTB1_STRPN]I6L8Q3 32.35 12 340 114.44 Choline binding protein F N- OS =Streptococcus pneumoniae terminally serotype 4 (strain ATCC BAA-334/anchored TIGR4) GN = cbpF PE = 4 SV = 1 − (with CS) [I6L8Q3_STRPN]Q97RS8 62.96 16 378 110.09 Lactate oxidase OS = StreptococcusIntracellular pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =lctO-2 PE = 4 SV = 1 − [Q97RS8_STRPN] Q97N53 70.96 11 272 109.3 Cellshape-determining protein MreC N- OS = Streptococcus pneumoniaeterminally serotype 4 (strain ATCC BAA-334/ anchored TIGR4) GN = mreC PE= 3 SV = 1 − (No CS) [Q97N53_STRPN] P0A3M5 41.32 18 680 109.25Penicillin-binding protein 2B N- OS = Streptococcus pneumoniaeterminally serotype 4 (strain ATCC BAA-334/ anchored TIGR4) GN = penA PE= 3 SV = 1 − (No CS) [PBP2_STRPN] P0A4S1 49.15 9 293 109Fructose-bisphosphate aldolase Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = fba PE = 3 SV =1 − [ALF_STRPN] Q97RK0 29.57 13 575 107.28 Septation ring formationregulator N- EzrA OS = Streptococcus pneumoniae terminally serotype 4(strain ATCC BAA-334/ anchored TIGR4) GN = ezrA PE = 3 SV = 1 − (No CS)[EZRA_STRPN] Q97SI9 60.55 7 109 107.21 Cell cycle protein GpsBIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = gpsB PE = 3 SV = 1 − [GPSB_STRPN] P63413 36.36 12396 106.75 Acetate kinase OS = Streptococcus Intracellular pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = ackA PE = 3 SV = 1 −[ACKA_STRPN] P0A2Z2 59.76 10 164 105.18 ATP synthase subunit bIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = atpF PE = 3 SV = 1 − [ATPF_STRPN] Q97SQ4 39.74 8156 103.23 30S ribosomal protein S7 Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = rpsG PE = 3 SV =1 − [RS7_STRPN] Q97QK5 48.6 19 607 101.3 Elongation factor 4 OS =Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = lepA PE = 3 SV = 1 − [LEPA_STRPN] P65239 39.75 11322 100.54 Ribose-phosphate pyrophosphokinase Intracellular 1 OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =prs1 PE = 3 SV = 1 − [KPRS1_STRPN] P95829 36.41 14 607 100.51 Chaperoneprotein DnaK Intracellular OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = dnaK PE = 3 SV = 2 − [DNAK_STRPN]I6L8U5 55.47 11 411 100.01 3-oxoacyl-[acyl-carrier-protein]Intracellular synthase 2 OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = fabF PE = 3 SV = 1 − [I6L8U5_STRPN]Q97PV1 21.8 6 289 93.98 Putative glycerol uptake facilitator Multi-protein OS = Streptococcus trans- pneumoniae serotype 4 (strain ATCCmembrane BAA-334/TIGR4) GN = SP_1491 PE = 3 SV = 1 − [Q97PV1_STRPN]Q2MGH6 15.73 18 1767 93.68 Endo-alpha-N-acetylgalactosaminidase LPxTG OS= Streptococcus pneumoniae Cell-wall serotype 4 (strain ATCC BAA-334/anchored TIGR4) GN = SP_0368 PE = 1 SV = 1 − [GH101_STRPN] Q97SQ9 36.0513 602 92.56 Glutamine-fructose-6-phosphate Intracellularaminotransferase [isomerizing] OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = glmS PE = 3 SV = 3 − [GLMS_STRPN]Q97P40 53.96 9 202 92.31 Putative general stress protein 24Intracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = SP_1804 PE = 4 SV = 1 − [Q97P40_STRPN] Q97QE4 25.3318 1058 90.95 Carbamoyl-phosphate synthase large Intracellular chain OS= Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =carB PE = 3 SV = 1 − [CARB_STRPN] I6L8Q9 40.04 11 457 90.76 Celldivision protein ftsA Intracellular OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = ftsA PE = 3 SV = 1 −[I6L8Q9_STRPN] I6L8S9 45.52 10 413 90.26 D-alanyl-D-alaninecarboxypeptidase N- OS = Streptococcus pneumoniae terminally serotype 4(strain ATCC BAA-334/ anchored TIGR4) GN = dacA PE = 3 SV = 1 − (withCS) [I6L8S9_STRPN] Q97Q48 27.52 18 872 89.81 Alanine-tRNA ligaseIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = alaS PE = 3 SV = 1 − [SYA_STRPN] Q97QD5 64.52 12186 89.81 LemA protein OS = Streptococcus N- pneumoniae serotype 4(strain ATCC terminally BAA-334/TIGR4) GN = lemA PE = 1 SV = 1 −anchored [Q97QD5_STRPN] (No CS) Q97NW2 48.69 13 419 88.69 Sugar ABCtransporter, sugar-binding Lipid protein OS = Streptococcus anchoredpneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = rafE PE = 1 SV =1 − [Q97NW2_STRPN] I6L8N0 43.01 14 365 88.6 UDP-N-acetylglucosamine-2-Intracellular epimerase OS = Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/TIGR4) GN = cps4I PE = 3 SV = 1 − [I6L8N0_STRPN] Q97QW436.47 13 425 87.93 Uncharacterized protein Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =SP_1083 PE = 4 SV = 1 − [Q97QW4_STRPN] Q9FBB7 58.43 12 255 86.89Acetyl-coenzyme A carboxylase Intracellular carboxyl transferase subunitalpha OS = Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = accA PE = 1 SV = 1 − [ACCA_STRPN] P67266 42.42 4 99 86.55Nucleoid-associated protein SP_1102 Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_1102 PE = 1SV = 1 − [Y1102_STRPN] Q97QM4 24.06 13 719 86.48Ribonucleoside-diphosphate Intracellular reductase OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = nrdE PE = 3 SV =1 − [Q97QM4_STRPN] Q97PF5 44.17 8 283 85.34 Putativephosphosugar-binding Intracellular transcriptional regulator OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =SP_1674 PE = 4 SV = 1 − [Q97PF5_STRPN] P0A3Y3 50.87 10 230 84.952,3-bisphosphoglycerate-dependent Intracellular phosphoglycerate mutaseOS = Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN= gpmA PE = 3 SV = 1 − [GPMA_STRPN] Q97NE1 40.78 4 103 84.35Uncharacterized protein Intracellular OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_2102 PE = 4 SV = 1 −[Q97NE1_STRPN] P67282 35.58 16 534 83.04 Ribonuclease Y OS =Streptococcus N- pneumoniae serotype 4 (strain ATCC terminallyBAA-334/TIGR4) GN = rny PE = 3 SV = 1 − anchored [RNY_STRPN] (No CS)Q97QX6 22.61 14 898 82.38 Phosphoenolpyruvate carboxylase IntracellularOS = Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN= ppc PE = 3 SV = 1 − [CAPP_STRPN] Q97SD1 32.63 12 521 81.72 Amino acidABC transporter, amino Multi- acid-binding protein/permease trans-protein OS = Streptococcus membrane pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_0453 PE = 4 SV = 1 − [Q97SD1_STRPN] P66419 50.568 89 78.12 30S ribosomal protein S14 Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = rpsN PE = 3 SV =1 − [RS14_STRPN] P61182 61.4 8 114 77.58 50S ribosomal protein L22Intracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = rplV PE = 1 SV = 1 − [RL22_STRPN] P66278 33.33 4 6677.34 50S ribosomal protein L35 Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = rpml PE = 3 SV =1 − [RL35_STRPN] Q97R14 34.57 10 324 76.27 Uncharacterized proteinSecretory OS = Streptococcus pneumoniae (released) serotype 4 (strainATCC BAA-334/ (with CS) TIGR4) GN = SP_1027 PE = 4 SV = 1 −[Q97R14_STRPN] Q97T52 22.4 12 616 70.86 Putative capsular polysaccharideMulti- biosynthesis protein trans- OS = Streptococcus pneumoniaemembrane serotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_0103 PE = 4 SV= 1 − [Q97T52_STRPN] Q97SW2 26.53 13 735 70.68 Anaerobic ribonucleoside-Intracellular triphosphate reductase OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = nrdD PE = 4 SV = 1 −[Q97SW2_STRPN] Q97SE4 32.3 12 514 70.62 Peptide chain release factor 3Intracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = prfC PE = 3 SV = 1 − [RF3_STRPN] Q97PC4 27.24 9 49268.5 ABC transporter, ATP-binding protein Multi- OS = Streptococcuspneumoniae trans- serotype 4 (strain ATCC BAA-334/ membrane TIGR4) GN =SP_1715 PE = 4 SV = 1 − [Q97PC4_STRPN] I6L8V8 47.33 7 243 68.483-oxoacyl-[acyl-carrier protein] Intracellular reductase OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =fabG PE = 3 SV = 1 − [I6L8V8_STRPN] Q97S28 21.17 11 737 68.26Polyribonucleotide Intracellular nucleotidyltransferase OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =pnp PE = 3 SV = 1 − [PNP_STRPN] Q97RC4 41.38 9 290 67.69 Uncharacterizedprotein Lipid OS = Streptococcus pneumoniae anchored serotype 4 (strainATCC BAA-334/ TIGR4) GN = SP_0899 PE = 4 SV = 1 − [Q97RC4_STRPN] Q97S8621.65 7 448 65.38 Glutamine synthetase, type I Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =glnA PE = 3 SV = 1 − [Q97S86_STRPN] Q97NH4 41.48 9 352 65.13 Alcoholdehydrogenase, zinc- Intracellular containing OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_2055 PE = 3 SV= 1 − [Q97NH4_STRPN] P0A495 65.79 3 38 64.65 50S ribosomal protein L36Intracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = rpmJ PE = 3 SV = 1 − [RL36_STRPN] Q97ND6 25.72 13587 64.48 Aspartate-tRNA ligase Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = aspS PE = 3 SV =1 − [SYD_STRPN] P72525 18.1 10 823 63.42 DNA topoisomerase 4 subunit AIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = parC PE = 1 SV = 3 − [PARC_STRPN] Q97PH2 23.11 12740 63.08 GTP pyrophosphokinase Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = relA PE = 3 SV =1 − [Q97PH2_STRPN] Q97T46 32.63 4 95 63.06 Putative bacteriocin OS =Streptococcus Secretory pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_0109 PE = 4 SV = 1 − [Q97T46_STRPN] I6L8V0 42.1711 351 60.21 Capsular polysaccharide biosynthesis Intracellular proteinCps4J OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = cap4J PE = 4 SV = 1 − [I6L8V0_STRPN] Q97RQ6 23.84 6281 60.06 DegV domain-containing protein Intracellular SP_0742 OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =SP_0742 PE = 3 SV = 1 − [Y742_STRPN] Q97SR2 35.08 9 419 60.03 Putativezinc metalloprotease Multi- SP_0263 OS = Streptococcus trans- pneumoniaeserotype 4 (strain ATCC membrane BAA-334/TIGR4) GN = SP_0263 PE = 3 SV =1 − [Y263_STRPN] Q97PB8 15.63 8 627 59.73 PTS system IIABC componentsMulti- OS = Streptococcus pneumoniae trans- serotype 4 (strain ATCCBAA-334/ membrane TIGR4) GN = SP_1722 PE = 4 SV = 1 − [Q97PB8_STRPN]Q97QE6 41.47 9 340 58.95 Alcohol dehydrogenase, zinc- Intracellularcontaining OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_1270 PE = 4 SV = 1 − [Q97QE6_STRPN] Q97P07 34.5610 408 58.93 Putative capsular polysaccharide Intracellular biosynthesisprotein OS = Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1837 PE = 3 SV = 1 − [Q97P07_STRPN] Q97T09 40.23 9 35357.99 Methionine import ATP-binding Intracellular protein MetN OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =metN PE = 3 SV = 1 − [METN_STRPN] Q97NP5 18.83 7 308 57.43 Membraneprotein insertase YidC 1 Multi- OS = Streptococcus pneumoniae trans-serotype 4 (strain ATCC BAA-334/ membrane TIGR4) GN = yidC1 PE = 3 SV =1 − [YIDC1_STRPN] P0A451 41.49 12 388 57.14 Protein RecA OS =Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = recA PE = 2 SV = 1 − [RECA_STRPN] Q97SU7 51.12 5 17856.93 50S ribosomal protein L6 Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = rplF PE = 3 SV =1 − [RL6_STRPN] Q97NE2 18.49 9 687 56.54 Cation-transporting ATPase,EI-E2 Intracellular family OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = SP_2101 PE = 3 SV = 1 − [Q97NE2_STRPN]Q97SN4 32.31 6 130 56.22 30S ribosomal protein S9 Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =rpsI PE = 3 SV = 1 − [RS9_STRPN] P66392 51.24 7 121 56.15 30S ribosomalprotein S13 Intracellular OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = rpsM PE = 3 SV = 1 − [RS13_STRPN]Q97S10 23.51 11 553 55.98 Ribonuclease J OS = StreptococcusIntracellular pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = rnjPE = 3 SV = 1 − [Q97S10_STRPN] Q97PG0 20.99 5 262 55.94 Cell divisionprotein DivIVA Intracellular OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = divIVA PE = 4 SV = 1 − [Q97PG0_STRPN]P35597 17.61 9 778 55.84 Probable cation-transporting ATPase Multi- exp7OS = Streptococcus pneumoniae trans- serotype 4 (strain ATCC BAA-334/membrane TIGR4) GN = exp7 PE = 3 SV = 2 − [EXP7_STRPN] Q97SJ2 43.52 12409 55.75 Capsular polysaccharide biosynthesis Intracellular proteinCps4K OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = cps4K PE = 4 SV = 1 − [Q97SJ2_STRPN] Q97PI4 29.98 12647 55.52 Threonine-tRNA ligase Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = thrS PE = 1 SV =1 − [SYT_STRPN] Q97QW8 25.81 8 434 55.41 GTPase Obg OS = StreptococcusIntracellular pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = obgPE = 3 SV = 1 − [OBG_STRPN] P67293 43.9 3 82 55.21 UPF0154 proteinSP_1882 N- OS = Streptococcus pneumoniae terminally serotype 4 (strainATCC BAA-334/ anchored TIGR4) GN = SP_1882 PE = 3 SV = 1 − (No CS)[Y1882_STRPN] P0A4L9 20.52 10 648 54.97 DNA gyrase subunit BIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = gyrB PE = 3 SV = 1 − [GYRB_STRPN] Q97SJ8 33.2 11494 54.78 UPF0371 protein SP_0341 Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_0341 PE = 3SV = 1 − [Y341_STRPN] Q54869 24.16 10 563 54.43 Arginine-tRNA ligaseIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = argS PE = 1 SV = 2 − [SYR_STRPN] P35595 14.88 7 72653.64 PTS system glucose-specific EIICBA Multi- component OS =Streptococcus trans- pneumoniae serotype 4 (strain ATCC membraneBAA-334/TIGR4) GN = exp5 PE = 3 SV = 2 − [PTG3C_STRPN] Q97SI7 22.41 7464 53.36 Mid-cell-anchored protein Z Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = mapZ PE = 1 SV =1 − [Q97SI7_STRPN] P0A4B5 55.91 7 93 52.72 30S ribosomal protein S19Intracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = rpsS PE = 3 SV = 1 − [RS19_STRPN] Q97SU3 23.29 4146 52.09 50S ribosomal protein L15 Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = rplO PE = 3 SV =1 − [RL15_STRPN] Q97RV5 27.57 11 613 51.74 Elongation factor Tu familyprotein Intracellular OS = Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/ TIGR4) GN = SP_0681 PE = 4 SV = 1 − [Q97RV5_STRPN] Q97NW98.4 5 655 50.97 Trehalose PTS system, IIABC Multi- components OS =Streptococcus trans- pneumoniae serotype 4 (strain ATCC membraneBAA-334/TIGR4) GN = SP_1884 PE = 4 SV = 1 − [Q97NW9_STRPN] Q97QE7 41.6 9262 49.97 Choline kinase OS = Streptococcus Intracellular pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = pck PE = 4 SV = 1 −[Q97QE7_STRPN] P67595 31.96 8 341 48.09 Tryptophan-tRNA ligaseIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = trpS PE = 3 SV = 1 − [SYW_STRPN] P66581 55.49 6 16447.77 30S ribosomal protein S5 Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = rpsE PE = 3 SV =1 − [RS5_STRPN] Q97R57 16.33 10 784 47.19 Ribonuclease R OS =Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = rnr PE = 3 SV = 1 − [Q97R57_STRPN] Q97RP0 21.03 7447 46.86 DEAD-box ATP-dependent RNA Intracellular helicase CshB OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =cshB PE = 3 SV = 1 − [Q97RP0_STRPN] P0A3R1 22.65 9 649 46.48 DNAmismatch repair protein HexB Intracellular OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = hexB PE = 3 SV = 1 −[HEXB_STRPN] P0A3R3 18.48 11 844 45.87 DNA mismatch repair protein HexAIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = hexA PE = 3 SV = 1 − [HEXA_STRPN] Q97SF6 27.97 7261 45.73 Enoyl-CoA hydratase/isomerase family Intracellular protein OS= Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =SP_0415 PE = 4 SV = 1 − [Q97SF6_STRPN] P66708 45.98 9 311 45.6DNA-directed RNA polymerase subunit Intracellular alpha OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =rpoA PE = 3 SV = 1 − [RPOA_STRPN] I6L8S8 28.69 9 481 45.33 Capsularpolysaccharide biosynthesis Multi- protein Cps4A OS = Streptococcustrans- pneumoniae serotype 4 (strain ATCC membrane BAA-334/TIGR4) GN =cps4A PE = 4 SV = 1 − [I6L8S8_STRPN] Q97N43 19.51 7 492 45.08Inosine-5′-monophosphate Intracellular dehydrogenase OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = guaB PE = 3 SV =1 − [Q97N43_STRPN] I6L8W8 16.87 6 332 44.57 Choline binding protein J N-OS = Streptococcus pneumoniae terminally serotype 4 (strain ATCCBAA-334/ anchored TIGR4) GN = cbpJ PE = 4 SV = 1 − (with CS)[I6L8W8_STRPN] Q2MGG2 27.62 7 467 44.4 Serine protease, subtilase familySecretory OS = Streptococcus pneumoniae (released) serotype 4 (strainATCC BAA-334/ (with CS) TIGR4) GN = SP_1954 PE = 4 SV = 1 −[Q2MGG2_STRPN] Q97SP0 42.22 7 270 44.36 Cof family protein OS =Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_0286 PE = 4 SV = 1 − [Q97SP0_STRPN] Q97PG9 38.757 240 44.16 Manganese ABC transporter, ATP- Intracellular bindingprotein OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = psaB PE = 3 SV = 1 − [Q97PG9_STRPN] Q97T72 31.67 6221 44.14 Potassium uptake protein, Trk family N- OS = Streptococcuspneumoniae terminally serotype 4 (strain ATCC BAA-334/ anchored TIGR4)GN = SP_0079 PE = 4 SV = 1 − (No CS) [Q97T72_STRPN] Q97RS9 21.98 7 49644.07 Lysine-tRNA ligase OS = Streptococcus Intracellular pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = lysS PE = 1 SV = 2 −[SYK_STRPN] Q97PK5 27.22 2 158 43.99 Uncharacterized protein N- OS =Streptococcus pneumoniae terminally serotype 4 (strain ATCC BAA-334/anchored TIGR4) GN = SP_1604 PE = 4 SV = 1 − (No CS) [Q97PK5_STRPN]Q97RV8 38.89 6 126 43.74 Uncharacterized protein N- OS = Streptococcuspneumoniae terminally serotype 4 (strain ATCC BAA-334/ anchored TIGR4)GN = SP_0678 PE = 4 SV = 1 − (No CS) [Q97RV8_STRPN] P06653 26.1 6 31843.37 Autolysin OS = Streptococcus Intracellular pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = lytA PE = 1 SV = 2 − [ALYS_STRPN]P63791 27.8 8 410 42.54 ATP-dependent Clp protease ATP- Intracellularbinding subunit ClpX OS = Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/ TIGR4) GN = clpX PE = 3 SV = 1 − [CLPX_STRPN] Q97Q36 37.337 217 42.45 Phosphate-specific transport system Intracellular accessoryprotein PhoU OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = SP_1395 PE = 3 SV = 1 − [Q97Q36_STRPN] Q97R16 32.0610 418 42.38 Serine hydroxymethyltransferase Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =glyA PE = 3 SV = 1 − [GLYA_STRPN] Q97NM1 38.99 9 336 42.33 Catabolitecontrol protein A Intracellular OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = ccpA PE = 4 SV = 1 − [Q97NM1_STRPN]Q97PQ8 42.66 6 143 40.36 Uncharacterized protein Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =SP_1545 PE = 4 SV = 1 − [Q97PQ8_STRPN] Q97SU6 59.32 7 118 40.13 50Sribosomal protein L18 Intracellular OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = rplR PE = 3 SV = 1 −[RL18_STRPN] Q97RC9 30.77 3 156 40.01 Arginine repressor OS =Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = argR PE = 3 SV = 1 − [Q97RC9_STRPN] Q97RC5 25.15 8501 39.74 Pyruvate kinase OS = Streptococcus Intracellular pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = pyk PE = 3 SV = 1 −[Q97RC5_STRPN] Q97PM1 45.04 7 262 38.81 GTP-sensing transcriptionalpleiotropic Intracellular repressor CodY OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = codY PE = 1 SV = 1 −[CODY_STRPN] I6L8N6 28.91 5 211 38.64 Capsular polysaccharidebiosynthesis N- protein Cps4E OS = Streptococcus terminally pneumoniaeserotype 4 (strain ATCC anchored BAA-334/TIGR4) GN = cps4E PE = 4 (NoCS) SV = 1 − [I6L8N6_STRPN] Q97R36 38.38 7 185 38.61 Thioredoxin familyprotein Lipid OS = Streptococcus pneumoniae anchored serotype 4 (strainATCC BAA-334/ TIGR4) GN = SP_1000 PE = 1 SV = 1 − [Q97R36_STRPN] Q97S9317.76 7 535 37.74 CTP synthase OS = Streptococcus Intracellularpneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = pyrG PE = 3 SV =1 − [PYRG_STRPN] Q97QW1 20.71 9 763 37.7 DNA helicase OS = StreptococcusIntracellular pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =pcrA PE = 4 SV = 1 − [Q97QW1_STRPN] P63544 51.76 6 170 37.59 Adeninephosphoribosyltransferase Intracellular OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = apt PE = 1 SV = 1 −[APT_STRPN] P65241 18.18 5 319 37.57 Ribose-phosphate pyrophosphokinaseIntracellular 2 OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = prs2 PE = 3 SV = 1 − [KPRS2_STRPN] Q97RE5 17.08 6650 37.17 PTS system, fructose specific IIABC Multi- components OS =Streptococcus trans- pneumoniae serotype 4 (strain ATCC membraneBAA-334/TIGR4) GN = SP_0877 PE = 4 SV = 1 − [Q97RE5_STRPN] P35596 21.389 608 37.12 Alpha-glycerophosphate oxidase N- OS = Streptococcuspneumoniae terminally serotype 4 (strain ATCC BAA-334/ anchored TIGR4)GN = glpO PE = 3 SV = 2 − (No CS) [GLPO_STRPN] Q97R12 19.52 7 543 37.11Uncharacterized RNA Intracellular methyltransferase SP_1029 OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =SP_1029 PE = 3 SV = 1 − [Y1029_STRPN] P64166 12.91 7 767 36.81 DNAtranslocase FtsK Multi- OS = Streptococcus pneumoniae trans- serotype 4(strain ATCC BAA-334/ membrane TIGR4) GN = ftsK PE = 3 SV = 1 −[FTSK_STRPN] P35594 17.15 10 752 36.7 ATP-dependent Clp protease ATP-Intracellular binding subunit ClpE OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = clpE PE = 3 SV = 2 −[CLPE_STRPN] P64072 53.33 8 195 36.63 Probable GTP-binding protein EngBIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = engB PE = 3 SV = 1 − [ENGB_STRPN] Q97TC4 31.11 5180 36.56 Hypoxanthine-guanine Intracellular phosphoribosyltransferaseOS = Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN= hpt PE = 3 SV = 1 − [HPRT_STRPN] Q97PU2 37.8 4 209 36.45 Amino acidABC transporter, ATP- Intracellular binding protein OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1501 PE = 3 SV= 1 − [Q97PU2_STRPN] P0A3S3 38.69 6 274 36.43 DNA-entry nuclease OS =Streptococcus N- pneumoniae serotype 4 (strain ATCC terminallyBAA-334/TIGR4) GN = endA PE = 1 SV = 1 − anchored [NUCE_STRPN] (No CS)I6L8N9 25.49 8 506 36.2 Choline transporter OS = Streptococcus Multi-pneumoniae serotype 4 (strain ATCC trans- BAA-334/TIGR4) GN = proWX PE =3 membrane SV = 1 − [I6L8N9_STRPN] Q97PQ5 25.99 8 531 36.19Uncharacterized protein Multi- OS = Streptococcus pneumoniae trans-serotype 4 (strain ATCC BAA-334/ membrane TIGR4) GN = SP_1548 PE = 4 SV= 1 − [Q97PQ5_STRPN] Q97NI7 28.66 6 328 36.13 Putative jag protein OS =Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_2040 PE = 4 SV = 1 − [Q97NI7_STRPN] P63742 19.327 502 36.12 Glycerol kinase OS = Streptococcus Intracellular pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = glpK PE = 3 SV = 1 −[GLPK_STRPN] Q97NT7 32.91 4 158 36 Uncharacterized protein Multi- OS =Streptococcus pneumoniae trans- serotype 4 (strain ATCC BAA-334/membrane TIGR4) GN = SP_1926 PE = 4 SV = 1 − [Q97NT7_STRPN] P64297 23.857 520 35.92 GMP synthase [glutamine-hydrolyzing] Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =guaA PE = 3 SV = 1 − [GUAA_STRPN] Q97R84 28.38 7 444 35.81Methylenetetrahydrofolate-tRNA- Intracellular(uracil-5-)-methyltransferase TrmFO OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = trmFO PE = 3 SV = 2 −[TRMFO_STRPN] Q97PK2 19.17 5 339 35.39 UDP-glucose 4-epimeraseIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = galE-1 PE = 3 SV = 1 − [Q97PK2_STRPN] Q97RP6 41.536 236 35.16 Branched-chain amino acid ABC Intracellular transporter,ATP-binding protein OS = Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/ TIGR4) GN = livF PE = 3 SV = 1 − [Q97RP6_STRPN] Q97NM330.74 10 462 35.12 Cof family protein OS = Streptococcus Intracellularpneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1997 PE = 4 SV= 1 − [Q97NM3_STRPN] Q97S34 10.86 7 801 34.77 Phenylalanine-tRNA ligasebeta Intracellular subunit OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = pheT PE = 3 SV = 1 − [SYFB_STRPN]Q97NQ4 31.38 9 427 34.05 UDP-N-acetylglucosamine 1- Intracellularcarboxyvinyltransferase 1 OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = murA1 PE = 3 SV = 1 − [MURA1_STRPN]Q97PY0 33.2 6 247 34.04 Amino acid ABC transporter, ATP- Intracellularbinding protein OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_1460 PE = 3 SV = 1 − [Q97PY0_STRPN] Q97RQ3 44.5 6209 33.91 Uracil phosphoribosyltransferase Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =upp PE = 3 SV = 1 − [UPP_STRPN] Q97QP0 15.87 7 567 33.86 Dihydrolipoyldehydrogenase Intracellular OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = SP_1161 PE = 4 SV = 1 − [Q97QP0_STRPN]P64062 24.77 8 436 33.49 GTPase Der OS = Streptococcus Intracellularpneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = der PE = 3 SV = 1− [DER_STRPN] Q97N48 22.83 6 276 33.26 Uncharacterized proteinIntracellular/ OS = Streptococcus pneumoniae TMH serotype 4 (strain ATCCBAA-334/ start TIGR4) GN = SP_2223 PE = 4 SV = 1 − AFTER 60[Q97N48_STRPN] Q97SJ5 22.98 7 409 32.72 Capsular polysaccharidebiosynthesis Multi- protein Cps4F OS = Streptococcus trans- pneumoniaeserotype 4 (strain ATCC membrane BAA-334/TIGR4) GN = cps4F PE = 4 SV = 1− [Q97SJ5_STRPN] P65832 36.73 6 275 32.69 Pur operon repressorIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = purR PE = 3 SV = 1 − [PURR_STRPN] P63384 12.09 9943 31.63 UvrABC system protein A Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = uvrA PE = 3 SV =1 − [UVRA_STRPN] I6L8W3 33.88 8 242 31.53 Choline transporter OS =Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = proV PE = 3 SV = 1 − [I6L8W3_STRPN] Q9AHD2 40.97 8227 31.44 Tyrosine-protein kinase CpsD Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = cpsD PE = 3 SV =1 − [CPSD_STRPN] O08397 23.84 8 453 31.17 Chromosomal replicationinitiator Intracellular protein DnaA OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = dnaA PE = 3 SV = 2 −[DNAA_STRPN] Q97PT5 23.63 7 292 30.59 ATP synthase gamma chainIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = atpG PE = 3 SV = 1 − [ATPG_STRPN] Q97PJ1 22.09 5249 30.29 Acyltransferase family protein Multi- OS = Streptococcuspneumoniae trans- serotype 4 (strain ATCC BAA-334/ membrane TIGR4) GN =SP_1624 PE = 4 SV = 1 − [Q97PJ1_STRPN] Q97NF6 14.36 6 564 30.07 ABCtransporter, ATP- Multi- binding/permease protein trans- OS =Streptococcus pneumoniae membrane serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_2075 PE = 3 SV = 1 − [Q97NF6_STRPN] Q97PW8 18.36 6 41430.06 Putative oxidoreductase Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_1472 PE = 4SV = 1 − [Q97PW8_STRPN] Q97QT0 15.61 5 474 29.86Glyceraldehyde-3-phosphate Intracellular dehydrogenase, NADP-dependentOS = Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN= gapN PE = 3 SV = 1 − [Q97QT0_STRPN] Q97PW6 12.24 6 678 29.5Glycine-tRNA ligase beta subunit Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = glyS PE = 3 SV =1 − [SYGB_STRPN] Q97T93 25.93 8 432 29.32 Adenylosuccinate lyaseIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = purB PE = 3 SV = 1 − [Q97T93_STRPN] Q97RG3 5.68 2229 29.15 Putative membrane protein Multi- OS = Streptococcus pneumoniaetrans- serotype 4 (strain ATCC BAA-334/ membrane TIGR4) GN = SP_0858 PE= 4 SV = 1 − [Q97RG3_STRPN] Q97TA6 12.42 3 330 28.21 Phosphateacyltransferase Intracellular OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = plsX PE = 3 SV = 1 − [PLSX_STRPN]Q97TA8 19.02 6 389 27.91 Aromatic amino acid aminotransferaseIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = araT PE = 3 SV = 1 − [Q97TA8_STRPN] P0A3B7 23.99 5346 27.48 Elongation factor Ts OS = Streptococcus Intracellularpneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = tsf PE = 1 SV = 2− [EFTS_STRPN] I6L8Q7 20.99 4 324 27.27 Enoyl-(Acyl-carrier-protein)reductase Intracellular OS = Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/ TIGR4) GN = fabK PE = 4 SV = 1 − [I6L8Q7_STRPN] Q97SG59.38 5 778 26.77 Endonuclease MutS2 Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = mutS2 PE = 3 SV= 1 − [MUTS2_STRPN] Q97S79 15.4 5 487 26.69 Type Irestriction-modification system, Intracellular M subunit OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =hsdM PE = 4 SV = 1 − [Q97S79_STRPN] Q97SF8 12.78 4 454 26.54Aspartokinase OS = Streptococcus Intracellular pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = SP_0413 PE = 3 SV = 1 − [Q97SF8_STRPN]Q97S89 19.1 5 398 26.46 Phosphoglycerate kinase Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =pgk PE = 3 SV = 1 − [PGK_STRPN] Q97QH0 22.14 6 429 26.44 Signalrecognition particle receptor Intracellular FtsY OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = ftsY PE = 1 SV =1 − [Q97QH0_STRPN] P0CB59 5.83 3 326 26.31 Phospho-N-acetylmuramoyl-Multi- pentapeptide-transferase trans- OS = Streptococcus pneumoniaemembrane serotype 4 (strain ATCC BAA-334/ TIGR4) GN = mraY PE = 3 SV = 1− [MRAY_STRPN] Q97P92 23.64 5 368 26.11 GTP-binding protein OS =Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_1749 PE = 4 SV = 1 − [Q97P92_STRPN] P63733 20.066 359 25.98 Carbamoyl-phosphate synthase small Intracellular chain OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =carA PE = 3 SV = 1 − [CARA_STRPN] Q97SP1 28.61 6 339 25.89 Alcoholdehydrogenase, zinc- Intracellular containing OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_0285 PE = 3 SV= 1 − [Q97SP1_STRPN] Q97PP2 19.92 4 256 25.75 Uncharacterized proteinMulti- OS = Streptococcus pneumoniae trans- serotype 4 (strain ATCCBAA-334/ membrane TIGR4) GN = SP_1561 PE = 1 SV = 1 − [Q97PP2_STRPN]P66339 48.04 4 102 25.74 30S ribosomal protein S10 Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =rpsJ PE = 3 SV = 1 − [RS10_STRPN] P0A335 13.52 6 540 25.62 60 kDachaperonin OS = Streptococcus Intracellular pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = groL PE = 1 SV = 1 − [CH60_STRPN]O07344 15.69 3 204 25.56 Signal peptidase I OS = Streptococcus N-pneumoniae serotype 4 (strain ATCC terminally BAA-334/TIGR4) GN = lepBPE = 3 SV = 2 − anchored [LEP_STRPN] (No CS) Q97PP5 41.41 3 128 25.55Uncharacterized protein Intracellular OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_1558 PE = 1 SV = 1 −[Q97PP5_STRPN] Q97RP7 15.75 2 254 25.33 Branched-chain amino acid ABCIntracellular transporter, ATP-binding protein OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = livG PE = 3 SV =1 − [Q97RP7_STRPN] Q97Q12 13.37 5 486 24.99 Nicotinatephosphoribosyltransferase Intracellular OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_1421 PE = 3 SV = 1 −[Q97Q12_STRPN] P67506 26.07 5 211 24.79 tRNA (guanine-N(7)-)-Intracellular methyltransferase OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = trmB PE = 1 SV = 1 − [TRMB_STRPN]P66155 17.74 2 62 24.21 50S ribosomal protein L28 Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =rpmB PE = 3 SV = 1 − [RL28_STRPN] P66200 71.25 3 80 23.95 50S ribosomalprotein L31 type B Intracellular OS = Streptococcus pneumoniae serotype4 (strain ATCC BAA-334/ TIGR4) GN = rpmE2 PE = 3 SV = 1 − [RL31B_STRPN]Q97NH2 17.69 5 605 23.83 Uncharacterized protein Multi- OS =Streptococcus pneumoniae trans- serotype 4 (strain ATCC BAA-334/membrane TIGR4) GN = SP_2057 PE = 4 SV = 1 − [Q97NH2_STRPN] Q97PP4 12.894 450 23.67 Phosphoglucosamine mutase Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = glmM PE = 3 SV =1 − [GLMM_STRPN] Q97R44 50.43 4 115 23.65 Uncharacterized proteinIntracellular/ OS = Streptococcus pneumoniae TMH serotype 4 (strain ATCCBAA-334/ start TIGR4) GN = SP_0990 PE = 4 SV = 1 − AFTER 60[Q97R44_STRPN] Q97QB4 15.18 4 448 23.57 Glutamate dehydrogenaseIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = gdhA PE = 3 SV = 1 − [Q97QB4_STRPN] Q97SW6 23.68 2152 23.55 Uncharacterized protein Lipid OS = Streptococcus pneumoniaeanchored serotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_0198 PE = 1 SV= 1 − [Q97SW6_STRPN] Q97PW5 16.07 4 305 23.51 Glycine-tRNA ligase alphasubunit Intracellular OS = Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/ TIGR4) GN = glyQ PE = 3 SV = 1 − [SYGA_STRPN] Q97Q69 19.635 428 23.47 Homoserine dehydrogenase Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = hom PE = 3 SV =1 − [Q97Q69_STRPN] Q97QT2 12.27 6 652 23.37 DNA ligase OS =Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = ligA PE = 3 SV = 1 − [DNLI_STRPN] P0A4B3 33.72 3 8623.26 30S ribosomal protein S17 Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = rpsQ PE = 3 SV =1 − [RS17_STRPN] Q97NG0 14.92 4 449 23.07 Glucose-6-phosphate isomeraseIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = pgi PE = 3 SV = 1 − [G6PI_STRPN] Q97T58 14.41 3 35422.73 Conserved domain protein Multi- OS = Streptococcus pneumoniaetrans- serotype 4 (strain ATCC BAA-334/ membrane TIGR4) GN = SP_0097 PE= 4 SV = 1 − [Q97T58_STRPN] Q97P32 16.46 4 407 22.04 Tryptophan synthasebeta chain Intracellular OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = trpB PE = 3 SV = 1 − [TRPB_STRPN]Q97QX3 28.57 5 252 21.94 ABC transporter, ATP-binding proteinIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = SP_1071 PE = 3 SV = 1 − [Q97QX3_STRPN] Q97ST6 32.032 128 21.84 50S ribosomal protein L17 Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = rplQ PE = 3 SV =1 − [RL17_STRPN] Q97RB6 8.74 5 709 21.38 Putative transcriptionalregulator Intracellular OS = Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/ TIGR4) GN = SP_0908 PE = 4 SV = 1 − [Q97RB6_STRPN] Q97NB917.91 4 335 21.02 Uncharacterized protein Multi- OS = Streptococcuspneumoniae trans- serotype 4 (strain ATCC BAA-334/ membrane TIGR4) GN =SP_2132 PE = 4 SV = 1 − [Q97NB9_STRPN] P66524 37.93 3 58 20.52 30Sribosomal protein S21 Intracellular OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = rpsU PE = 3 SV = 1 −[RS21_STRPN] Q97SI6 14.98 5 474 20.22 6-phosphogluconate dehydrogenase,Intracellular decarboxylating OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = gnd PE = 3 SV = 1 − [Q97SI6_STRPN]Q97R47 30.45 4 266 20.09 Uncharacterized protein N- OS = Streptococcuspneumoniae terminally serotype 4 (strain ATCC BAA-334/ anchored TIGR4)GN = SP_0987 PE = 1 SV = 1 − (No CS) [Q97R47_STRPN]

TABLE 4 Proteins identified in MP from TIGR4. The accession numbersrefer to SwissProt on the date of filing. Accession # Coverage #Peptides #AAs Score Description Localization Q97RH0 76.86 46 350 2085.84Lipoprotein OS = Streptococcus Lipid pneumoniae serotype 4 (strain ATCCanchored BAA-334/TIGR4) GN = SP_0845 PE = 4 SV = 1 - [Q97RH0_STRPN]Q97PE6 90.27 56 442 1745.9 Sugar ABC transporter, sugar-binding Lipidprotein OS = Streptococcus pneumoniae anchored serotype 4 (strain ATCCBAA-334/ TIGR4) GN = SP_1683 PE = 4 SV = 1 - [Q97PE6_STRPN] Q97T39 61.2950 744 1569.47 Pneumococcal surface protein A N- OS = Streptococcuspneumoniae terminally serotype 4 (strain ATCC BAA-334/ anchored TIGR4)GN = pspA PE = 4 SV = 1 - (with CS) [Q97T39_STRPN] P59213 82.74 62 4231500.08 Maltose/maltodextrin-binding protein Lipid OS = Streptococcuspneumoniae anchored serotype 4 (strain ATCC BAA-334/ TIGR4) GN = malX PE= 1 SV = 1 - [MALX_STRPN] Q97N74 53.25 47 693 1294.91 Choline bindingprotein A N- OS = Streptococcus pneumoniae terminally serotype 4 (strainATCC BAA-334/ anchored TIGR4) GN = cbpA PE = 1 SV = 1 - (with CS)[Q97N74_STRPN] Q97R51 87.86 45 313 1201.93 Foldase protein PrsA Lipid OS= Streptococcus pneumoniae anchored serotype 4 (strain ATCC BAA-334/TIGR4) GN = prsA PE = 3 SV = 1 - [PRSA_STRPN] Q54970 70.05 54 5911000.77 Pyruvate oxidase OS = Streptococcus Intracellular pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = spxB PE = 3 SV = 2 -[POXB_STRPN] Q2MGF6 75.51 50 490 957.09 Lysozyme OS = StreptococcusSecretory pneumoniae serotype 4 (strain ATCC (released) BAA-334/TIGR4)GN = lytC PE = 4 SV = 1 - (with CS) [Q2MGF6_STRPN] Q97PT1 77.13 52 551947.11 Uncharacterized protein Intracellular/ OS = Streptococcuspneumoniae TMH serotype 4 (strain ATCC BAA-334/ start TIGR4) GN =SP_1518 PE = 4 SV = 1 - AFTER 60 [Q97PT1_STRPN] Q97QS2 75.35 31 434783.92 Enolase OS = Streptococcus Intracellular pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = eno PE = 1 SV = 1 - [ENO_STRPN] P6402276.77 48 693 749.91 Elongation factor G OS = Streptococcus Intracellularpneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = fusA PE = 3 SV =1 - [EFG_STRPN] Q04707 72.6 47 719 715.79 Penicillin-binding protein 1AN- OS = Streptococcus pneumoniae terminally serotype 4 (strain ATCCBAA-334/ anchored TIGR4) GN = ponA PE = 1 SV = 2 - (No CS) [PBPA_STRPN]P0C2J9 87.47 49 471 703.79 Pneumolysin OS = Streptococcus Intracellularpneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = ply PE = 3 SV =1 - [TACY_STRPN] Q97PE1 73.93 35 445 655.97 ABC transporter,substrate-binding Lipid protein OS = Streptococcus pneumoniae anchoredserotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_1690 PE = 4 SV = 1 -[Q97PE1_STRPN] Q97NQ8 61.8 68 1225 650.84 DNA-directed RNA polymerasesubunit Intracellular beta′ OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = rpoC PE = 3 SV = 1 - [RPOC_STRPN]Q97NL1 79.1 29 335 611.32 Glyceraldehyde-3-phosphate Intracellulardehydrogenase OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = gap PE = 3 SV = 1 - [Q97NL1_STRPN] Q97T12 82.97 33276 589.52 ABC transporter, substrate-binding Lipid protein OS =Streptococcus pneumoniae anchored serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_0148 PE = 4 SV = 1 - [Q97T12_STRPN] Q97T63 83.3 36 491576.45 ABC transporter, substrate-binding Lipid protein OS =Streptococcus pneumoniae anchored serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_0092 PE = 4 SV = 1 - [Q97T63_STRPN] I6L8V7 44.02 33 627457.19 Choline binding protein E Secretory OS = Streptococcus pneumoniae(released) serotype 4 (strain ATCC BAA-334/ (with CS) TIGR4) GN = cbpEPE = 4 SV = 1 - [I6L8V7_STRPN] Q97NQ7 55.61 50 1203 449.03 DNA-directedRNA polymerase subunit Intracellular beta OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = rpoB PE = 3 SV = 1 -[RPOB_STRPN] Q97SV2 64.62 18 277 395.51 50S ribosomal protein L2Intracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = rplB PE = 3 SV = 1 - [RL2_STRPN] P59205 60.33 29658 388.39 Putative endo-beta-N- Secretory acetylglucosaminidase(released) OS = Streptococcus pneumoniae (with CS) serotype 4 (strainATCC BAA-334/ TIGR4) GN = lytB PE = 1 SV = 1 - [LYTB_STRPN] O69076 55.9831 652 385.64 ATP-dependent zinc metalloprotease Multi- FtsH OS =Streptococcus pneumoniae transmembrane serotype 4 (strain ATCC BAA-334/TIGR4) GN = ftsH PE = 3 SV = 3 - [FTSH_STRPN] Q97SP2 67.47 25 332 369.57PTS system, mannose-specific IIAB Intracellular components OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =manL PE = 4 SV = 1 - [Q97SP2_STRPN] Q97T11 66.55 23 284 361.41Lipoprotein OS = Streptococcus Lipid pneumoniae serotype 4 (strain ATCCanchored BAA-334/TIGR4) GN = SP_0149 PE = 3 SV = 1 - [Q97T11_STRPN]I6L8Q3 52.65 17 340 354.19 Choline binding protein F N- OS =Streptococcus pneumoniae terminally serotype 4 (strain ATCC BAA-334/anchored TIGR4) GN = cbpF PE = 4 SV = 1 - (with CS) [I6L8Q3_STRPN]Q97NX9 80.69 25 321 336.18 Iron-compound ABC transporter, iron- Lipidcompound-binding protein anchored OS = Streptococcus pneumoniae serotype4 (strain ATCC BAA-334/ TIGR4) GN = SP_1872 PE = 4 SV = 1 -[Q97NX9_STRPN] P64030 57.29 25 398 336.06 Elongation factor Tu OS =Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = tuf PE = 3 SV = 1 - [EFTU_STRPN] Q97NW2 64.44 25 419331.55 Sugar ABC transporter, sugar-binding Lipid protein OS =Streptococcus pneumoniae anchored serotype 4 (strain ATCC BAA-334/TIGR4) GN = rafE PE = 1 SV = 1 - [Q97NW2_STRPN] Q97QH2 42.86 29 721330.61 Amino acid ABC transporter, amino Multi- acid-bindingprotein/permease protein transmembrane OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_1241 PE = 1 SV = 1 -[Q97QH2_STRPN] Q97RS8 78.31 29 378 328.02 Lactate oxidase OS =Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = lctO-2 PE = 4 SV = 1 - [Q97R58_STRPN] Q97PW1 63.7125 463 316.66 Peptidoglycan N-acetylglucosamine N- deacetylase A OS =Streptococcus terminally pneumoniae serotype 4 (strain ATCC anchoredBAA-334/TIGR4) GN = pgdA PE = 4 SV = 1 - (No CS) [Q97PW1_STRPN] Q97PT675.85 26 468 315.76 ATP synthase subunit beta Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =atpD PE = 3 SV = 1 - [ATPB_STRPN] Q97NQ3 58.55 22 345 310Uncharacterized protein N- OS = Streptococcus pneumoniae terminallyserotype 4 (strain ATCC BAA-334/ anchored TIGR4) GN = SP_1967 PE = 4 SV= 1 - (No CS) [Q97NQ3_STRPN] Q97R09 58.94 17 341 309.62 Iron-compoundABC transporter, iron Lipid compound-binding protein anchored OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =SP_1032 PE = 1 SV = 1 - [Q97R09_STRPN] P18791 69.8 28 659 298.67Oligopeptide-binding protein AmiA Lipid OS = Streptococcus pneumoniaeanchored serotype 4 (strain ATCC BAA-334/ TIGR4) GN = amiA PE = 1 SV =3 - [AMIA_STRPN] Q97RZ7 63.03 15 238 283.45 Uncharacterized proteinLipid OS = Streptococcus pneumoniae anchored serotype 4 (strain ATCCBAA-334/ TIGR4) GN = SP_0629 PE = 1 SV = 1 - [Q97RZ7_STRPN] P0A4G2 65.0518 309 281.58 Manganese ABC transporter substrate- Lipid bindinglipoprotein OS = Streptococcus anchored pneumoniae serotype 4 (strainATCC BAA-334/TIGR4) GN = psaA PE = 1 SV = 1 - [MTSA_STRPN] P66112 60.510 119 280.56 50S ribosomal protein L20 Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = rplT PE = 3 SV =1 - [RL20_STRPN] Q97PM5 63.3 23 376 278.8 Sugar ABC transporter,ATP-binding Intracellular protein OS = Streptococcus pneumoniae serotype4 (strain ATCC BAA-334/ TIGR4) GN = msmK PE = 4 SV = 1 - [Q97PM5_STRPN]P0A4M7 41.57 23 498 271.04 Oligopeptide transport system Multi- permeaseprotein AmiC transmembrane OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = amiC PE = 3 SV = 1 - [AMIC_STRPN]Q97SP4 48.18 12 303 262.56 PTS system, mannose-specific IID Multi-component OS = Streptococcus transmembrane pneumoniae serotype 4 (strainATCC BAA-334/TIGR4) GN = SP_0282 PE = 4 SV = 1 - [Q97SP4_STRPN] Q97NL360.74 34 731 258.29 Penicillin-binding protein 2A N- OS = Streptococcuspneumoniae terminally serotype 4 (strain ATCC BAA-334/ anchored TIGR4)GN = pbp2A PE = 4 SV = 1 - (No CS) [Q97NL3_STRPN] P14677 61.33 32 750257.72 Penicillin-binding protein 2x N- OS = Streptococcus pneumoniaeterminally serotype 4 (strain ATCC BAA-334/ anchored TIGR4) GN = pbpX PE= 1 SV = 2 - (No CS) [PBPX_STRPN] Q97PT4 38.92 21 501 249.37 ATPsynthase subunit alpha Multi- OS = Streptococcus pneumoniaetransmembrane serotype 4 (strain ATCC BAA-334/ TIGR4) GN = atpA PE = 3SV = 1 - [ATPA_STRPN] Q97NB5 52.66 30 621 236.54 Choline binding proteinPcpA Secretory OS = Streptococcus pneumoniae (released) serotype 4(strain ATCC BAA-334/ (with CS) TIGR4) GN = pcpA PE = 1 SV = 1 -[Q97NB5_STRPN] Q97PQ2 33.26 21 914 236.23 Cation-transporting ATPase,E1-E2 Multi- family OS = Streptococcus pneumoniae transmembrane serotype4 (strain ATCC BAA-334/ TIGR4) GN = SP_1551 PE = 3 SV = 1 -[Q97PQ2_STRPN] P0A475 50.36 8 137 229.6 50S ribosomal protein L16Intracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = rplP PE = 1 SV = 1 - [RL16_STRPN] Q97PU3 64.03 23278 227.7 Amino acid ABC transporter, amino Lipid acid-binding proteinOS = Streptococcus anchored pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = aatB PE = 4 SV = 1 - [Q97PU3_STRPN] Q97RC5 50.3 20501 223.13 Pyruvate kinase OS = Streptococcus Intracellular pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = pyk PE = 3 SV = 1 -[Q97RC5_STRPN] Q97NS5 65.68 19 338 221.3 Putative transcriptionalregulator Secretory OS = Streptococcus pneumoniae (released) serotype 4(strain ATCC BAA-334/ (with CS) TIGR4) GN = SP_1942 PE = 4 SV = 1 -[Q97NS5_STRPN] Q97RP4 75.22 14 230 213.93 Cell division ABC transporter,ATP- Intracellular binding protein FtsE OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = ftsE PE = 3 SV = 1 -[Q97RP4_STRPN] Q97RN2 66.67 13 267 207.19 Peptidyl-prolyl cis-transisomerase, Lipid cyclophilin-type OS = Streptococcus anchored pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_0771 PE = 3 SV = 1 -[Q97RN2_STRPN] P66565 61.58 21 203 205.82 30S ribosomal protein S4Intracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = rpsD PE = 3 SV = 1 - [RS4_STRPN] Q97QH1 61.79 10246 202.96 Amino acid ABC transporter, ATP- Intracellular bindingprotein OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_1242 PE = 3 SV = 1 - [Q97QH1_STRPN] I6L8N9 40.9118 506 201.9 Choline transporter OS = Streptococcus Multi- pneumoniaeserotype 4 (strain ATCC transmembrane BAA-334/TIGR4) GN = proWX PE = 3SV = 1 - [I6L8N9_STRPN] Q97N69 55.22 15 335 201.71 Putative ABCtransporter, substrate- Lipid binding protein OS = Streptococcusanchored pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_2197PE = 4 SV = 1 - [Q97N69_STRPN] P65887 53.97 23 428 199.01Adenylosuccinate synthetase Intracellular OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = purA PE = 1 SV = 1 -[PURA_STRPN] P18766 60.71 13 308 198.48 Oligopeptide transportATP-binding Intracellular protein AmiF OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = amiF PE = 3 SV = 2 -[AMIF_STRPN] Q97QB4 61.83 23 448 197.42 Glutamate dehydrogenaseIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = gdhA PE = 3 SV = 1 - [Q97QB4_STRPN] Q97SP5 55.18 19444 196.27 Aminopeptidase C OS = Streptococcus Intracellular pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = pepC PE = 4 SV = 1 -[Q97SP5_STRPN] Q97RP3 40.58 17 308 193.01 Cell division protein FtsXMulti- OS = Streptococcus pneumoniae transmembrane serotype 4 (strainATCC BAA-334/ TIGR4) GN = ftsX PE = 3 SV = 1 - [Q97RP3_STRPN] Q97T4636.84 5 95 189.78 Putative bacteriocin OS = Streptococcus Secretorypneumoniae serotype 4 (strain ATCC (released) BAA-334/TIGR4) GN =SP_0109 PE = 4 (no CS) SV = 1 - [Q97T46_STRPN] Q97PD6 51.85 31 837188.69 Protein translocase subunit SecA 1 Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =secA1 PE = 3 SV = 1 - [SECA1_STRPN] P0A4A7 60.58 13 137 187.87 30Sribosomal protein S12 Intracellular OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = rpsL PE = 3 SV = 1 -[RS12_STRPN] P0A2U8 52.96 20 355 185.88 Oligopeptide transportATP-binding Intracellular protein AmiE OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = amiE PE = 3 SV = 1 -[AMIE_STRPN] Q97RQ0 70.21 18 386 182.7 Branched-chain amino acid ABCLipid transporter, amino acid-binding anchored protein OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =livJ PE = 4 SV = 1 - [Q97RQ0_STRPN] P0A3M9 67.68 16 328 176.61 L-lactatedehydrogenase Intracellular OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = Idh PE = 3 SV = 2 - [LDH_STRPN] Q97RG949.51 20 511 174.4 Sugar ABC transporter, ATP-binding Intracellularprotein OS = Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_0846 PE = 3 SV = 1 - [Q97RG9_STRPN] P0A3M5 43.38 22 680166.84 Penicillin-binding protein 2B N- OS = Streptococcus pneumoniaeterminally serotype 4 (strain ATCC BAA-334/ anchored TIGR4) GN = penA PE= 3 SV = 1 - (No CS) [PBP2_STRPN] Q97N53 63.24 12 272 163.14 Cellshape-determining protein MreC N- OS = Streptococcus pneumoniaeterminally serotype 4 (strain ATCC BAA-334/ anchored TIGR4) GN = mreC PE= 3 SV = 1 - (No CS) [Q97N53_STRPN] P95829 49.26 20 607 161.66 Chaperoneprotein DnaK Intracellular OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = dnaK PE = 3 SV = 2 - [DNAK_STRPN]Q97SI9 67.89 7 109 156.35 Cell cycle protein GpsB Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =gpsB PE = 3 SV = 1 - [GPSB_STRPN] Q97NE4 36.3 26 821 153.09Penicillin-binding protein 1B N- OS = Streptococcus pneumoniaeterminally serotype 4 (strain ATCC BAA-334/ anchored TIGR4) GN = pbp1BPE = 4 SV = 1 - (No CS) [Q97NE4_STRPN] I6L8W3 71.49 15 242 152.01Choline transporter OS = Streptococcus Intracellular pneumoniae serotype4 (strain ATCC BAA-334/TIGR4) GN = proV PE = 3 SV = 1 - [I6L8W3_STRPN]P0A335 52.04 21 540 151.44 60 kDa chaperonin OS = StreptococcusIntracellular pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =groL PE = 1 SV = 1 - [CH60_STRPN] Q97NG0 44.77 17 449 145.04Glucose-6-phosphate isomerase Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = pgi PE = 3 SV =1 - [G6PI_STRPN] Q97Q31 57.53 12 292 143.85 Phosphate-binding proteinPstS 1 Lipid OS = Streptococcus pneumoniae anchored serotype 4 (strainATCC BAA-334/ TIGR4) GN = pstS1 PE = 1 SV = 1 - [PSTS1_STRPN] Q97RW951.81 16 332 139.95 Putative pneumococcal surface protein Lipid OS =Streptococcus pneumoniae anchored serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_0667 PE = 1 SV = 1 - [Q97RW9_STRPN] P22976 44.98 18 658139.63 Probable transketolase Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = tkt PE = 3 SV =2 - [TKT_STRPN] Q97SQ9 42.52 17 602 138.65Glutamine--fructose-6-phosphate Intracellular aminotransferase[isomerizing] OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = glmS PE = 3 SV = 3 - [GLMS_STRPN] Q97PA9 33.84 18659 132.79 Serine/threonine-protein kinase StkP Intracellular/ OS =Streptococcus pneumoniae TMH serotype 4 (strain ATCC BAA-334/ startTIGR4) GN = stkP PE = 1 SV = 1 - AFTER 60 [STKP2_STRPN] Q97Q34 70.41 18267 131.91 Phosphate import ATP-binding protein Intracellular PstB 2 OS= Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =pstB2 PE = 3 SV = 1 - [PSTB2_STRPN] Q97QX5 55.81 14 344 130.66Uncharacterized protein Secretory OS = Streptococcus pneumoniae(released) serotype 4 (strain ATCC BAA-334/ (with CS) TIGR4) GN =SP_1069 PE = 1 SV = 1 - [Q97QX5_STRPN] I6L8V0 55.84 18 351 130.62Capsular polysaccharide biosynthesis Intracellular protein Cps4J OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =cap4J PE = 4 SV = 1 - [I6L8V0_STRPN] Q97SD1 43.38 17 521 119.96 Aminoacid ABC transporter, amino Multi- acid-binding protein/permease proteintransmembrane OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = SP_0453 PE = 4 SV = 1 - [Q97SD1_STRPN] Q97SR1 50.7321 617 119.02 Proline--tRNA ligase OS = Streptococcus Intracellularpneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = proS PE = 3 SV =1 - [SYP_STRPN] P0A4D7 36.07 14 524 117.93 DEAD-box ATP-dependent RNAIntracellular helicase CshA OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = cshA PE = 3 SV = 1 - [CSHA_STRPN]Q97P40 68.81 12 202 117.92 Putative general stress protein 24Intracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = SP_1804 PE = 4 SV = 1 - [Q97P40_STRPN] Q97NK0 41 21883 117.4 Aldehyde-alcohol dehydrogenase Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =SP_2026 PE = 3 SV = 1 - [Q97NK0_STRPN] Q97N99 51.82 14 274 115.56 SPFHdomain/Band 7 family Intracellular OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_2156 PE = 4 SV = 1 -[Q97N99_STRPN] Q9L7Q2 17.63 23 1906 114.8 Zinc metalloprotease ZmpBMulti- OS = Streptococcus pneumoniae transmembrane serotype 4 (strainATCC BAA-334/ TIGR4) GN = zmpB PE = 3 SV = 2 - [ZMPB_STRPN] P67293 43.94 82 114.56 UPF0154 protein SP_1882 N- OS = Streptococcus pneumoniaeterminally serotype 4 (strain ATCC BAA-334/ anchored TIGR4) GN = SP_1882PE = 3 SV = 1 - (No CS) [Y1882_STRPN] Q97N37 61.83 14 393 114.16 Serineprotease OS = Streptococcus N- pneumoniae serotype 4 (strain ATCCterminally BAA-334/TIGR4) GN = SP_2239 PE = 1 anchored SV = 1 -[Q97N37_STRPN] (No CS) P66359 55.12 8 127 112.68 30S ribosomal proteinS11 Intracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = rpsK PE = 3 SV = 1 - [RS11_STRPN] Q97PX1 30.72 12459 110.91 NADH oxidase OS = Streptococcus Intracellular pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = nox PE = 4 SV = 1 -[Q97PX1_STRPN] P35592 43.64 21 660 109.44 Oligopeptide-binding proteinAliA Lipid OS = Streptococcus pneumoniae anchored serotype 4 (strainATCC BAA-334/ TIGR4) GN = aliA PE = 3 SV = 4 - [ALIA_STRPN] Q97PC4 28.6611 492 108.41 ABC transporter, ATP-binding protein Multi- OS =Streptococcus pneumoniae transmembrane serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1715 PE = 4 SV = 1 - [Q97PC4_STRPN] Q97P68 69.23 8 104104.87 Thioredoxin OS = Streptococcus Intracellular pneumoniae serotype4 (strain ATCC BAA-334/TIGR4) GN = trx PE = 3 SV = 1 - [Q97P68_STRPN]Q97PL8 41.55 12 438 101 Oxidoreductase, pyridine nucleotide-Intracellular disulfide, class I OS = Streptococcus pneumoniae serotype4 (strain ATCC BAA-334/TIGR4) GN = SP_1588 PE = 3 SV = 1 -[Q97PL8_STRPN] Q97SX2 68.78 17 189 97.17 Uncharacterized protein LipidOS = Streptococcus pneumoniae anchored serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_0191 PE = 1 SV = 1 - [Q97SX2_STRPN] P63413 43.94 14 39696.43 Acetate kinase OS = Streptococcus Intracellular pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = ackA PE = 3 SV = 1 -[ACKA_STRPN] Q97SJ6 52.61 11 230 96.34 Capsular polysaccharidebiosynthesis Multi- protein CpsC OS = Streptococcus transmembranepneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = cpsC PE = 3 SV =1 - [CPSC_STRPN] Q97Q67 38.08 11 344 95.87 Conserved domain proteinMulti- OS = Streptococcus pneumoniae transmembrane serotype 4 (strainATCC BAA-334/ TIGR4) GN = SP_1363 PE = 4 SV = 1 - [Q97Q67_STRPN] P0A4S149.15 10 293 93.48 Fructose-bisphosphate aldolase Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =fba PE = 3 SV = 1 - [ALF_STRPN] Q97SR2 37.71 11 419 93.31 Putative zincmetalloprotease SP_0263 Multi- OS = Streptococcus pneumoniaetransmembrane serotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_0263 PE =3 SV = 1 - [Y263_STRPN] Q97NP5 25.65 9 308 92.53 Membrane proteininsertase YidC 1 Multi- OS = Streptococcus pneumoniae transmembraneserotype 4 (strain ATCC BAA-334/ TIGR4) GN = yidC1 PE = 3 SV = 1 -[YIDC1_STRPN] Q97SJ4 44.44 15 360 91.77 Uncharacterized proteinIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = SP_0355 PE = 4 SV = 1 - [Q97SJ4_STRPN] Q97RK0 19.8310 575 91.07 Septation ring formation regulator N- EzrA OS =Streptococcus pneumoniae terminally serotype 4 (strain ATCC BAA-334/anchored TIGR4) GN = ezrA PE = 3 SV = 1 - (No CS) [EZRA_STRPN] P6337371.43 11 252 91.01 Phosphate import ATP-binding protein IntracellularPstB 1 OS = Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = pstB1 PE = 3 SV = 1 - [PSTB1_STRPN] Q97S89 51.01 13 39889.47 Phosphoglycerate kinase Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = pgk PE = 3 SV =1 - [PGK_STRPN] Q97PT5 47.26 12 292 85.91 ATP synthase gamma chainIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = atpG PE = 3 SV = 1 - [ATPG_STRPN] Q97SI4 32.65 9340 85.66 Choline binding protein C N- OS = Streptococcus pneumoniaeterminally serotype 4 (strain ATCC BAA-334/ anchored TIGR4) GN = cbpC PE= 4 SV = 1 - (with CS) [Q97SI4_STRPN] I6L8S8 39.71 14 481 85.62 Capsularpolysaccharide biosynthesis Multi- protein Cps4A OS = Streptococcustransmembrane pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =cps4A PE = 4 SV = 1 - [I6L8S8_STRPN] Q97PY0 65.59 13 247 85.07 Aminoacid ABC transporter, ATP- Intracellular binding protein OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =SP_1460 PE = 3 SV = 1 - [Q97PY0_STRPN] Q97SQ4 46.79 9 156 83.39 30Sribosomal protein S7 Intracellular OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = rpsG PE = 3 SV = 1 -[RS7_STRPN] P0A3Y3 60 10 230 82.54 2,3-bisphosphoglycerate-dependentIntracellular phosphoglycerate mutase OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = gpmA PE = 3 SV = 1 -[GPMA_STRPN] P0A4C3 59.91 11 217 82.05 30S ribosomal protein S3Intracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = rpsC PE = 3 SV = 1 - [RS3_STRPN] P65607 38.46 11338 79.74 Ornithine carbamoyltransferase, Intracellular catabolic OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN =arcB PE = 3 SV = 1 - [OTCC_STRPN] O05703 39.92 12 501 79.68 Zinc-bindinglipoprotein AdcA Lipid OS = Streptococcus pneumoniae anchored serotype 4(strain ATCC BAA-334/ TIGR4) GN = adcA PE = 3 SV = 4 - [ADCA_STRPN]I6L8U0 27.16 11 394 79.39 UDP-N-acetylglucosamine 2-epimeraseIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = cps4L PE = 3 SV = 1 - [I6L8U0_STRPN] Q97SI7 28.23 8464 78.19 Mid-cell-anchored protein Z Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = mapZ PE = 1 SV =1 - [Q97SI7_STRPN] Q97N56 58.3 15 259 78.07 30S ribosomal protein S2Intracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = rpsB PE = 3 SV = 1 - [RS2_STRPN] Q97SI6 35.44 13474 77.15 6-phosphogluconate dehydrogenase, Intracellulardecarboxylating OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = gnd PE = 3 SV = 1 - [Q97SI6_STRPN] Q97PG0 27.1 7 26277.07 Cell division protein DivIVA Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = divIVA PE = 4 SV= 1 - [Q97PG0_STRPN] Q97RF9 32 11 400 76.46 Ribosomal protein S1Intracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = rpsA PE = 4 SV = 1 - [Q97RF9_STRPN] Q97T80 15.68 171856 76.28 Zinc metalloprotease ZmpC LPxTG OS = Streptococcus pneumoniaeCell-wall serotype 4 (strain ATCC BAA-334/ anchored TIGR4) GN = zmpC PE= 3 SV = 1 - [ZMPC_STRPN] P0A2Z2 62.8 9 164 75.56 ATP synthase subunit bIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = atpF PE = 3 SV = 1 - [ATPF_STRPN] I6L8S9 47.7 10413 73.22 D-alanyl-D-alanine carboxypeptidase N- OS = Streptococcuspneumoniae terminally serotype 4 (strain ATCC BAA-334/ anchored TIGR4)GN = dacA PE = 3 SV = 1 - (with CS) [I6L8S9_STRPN] Q97PU4 80 7 115 71.8Bacterocin transport accessory protein Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = bta PE = 4 SV =1 - [Q97PU4_STRPN] Q97SU6 70.34 7 118 71.64 50S ribosomal protein L18Intracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = rplR PE = 3 SV = 1 - [RL18_STRPN] P63742 35.46 11502 71.21 Glycerol kinase OS = Streptococcus Intracellular pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = glpK PE = 3 SV = 1 -[GLPK_STRPN] Q97SN4 32.31 6 130 68.89 30S ribosomal protein S9Intracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = rpsI PE = 3 SV = 1 - [RS9_STRPN] P66907 31.84 6 38068.13 Queuine tRNA-ribosyltransferase Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = tgt PE = 3 SV =1 - [TGT_STRPN] Q97SF8 32.16 12 454 65.06 Aspartokinase OS =Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_0413 PE = 3 SV = 1 - [Q97SF8_STRPN] P0A471 82.799 122 62.07 50S ribosomal protein L7/L12 Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =rplL PE = 1 SV = 2 - [RL7_STRPN] I6L8W8 21.08 8 332 61.96 Cholinebinding protein J N- OS = Streptococcus pneumoniae terminally serotype 4(strain ATCC BAA-334/ anchored TIGR4) GN = cbpJ PE = 4 SV = 1 - (withCS) [I6L8W8_STRPN] P0A4J6 40.8 5 201 61.71 Superoxide dismutase [Mn]Intracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = sodA PE = 1 SV = 2 - [SODM_STRPN] Q97T52 30.03 14616 60.5 Putative capsular polysaccharide Multi- biosynthesis protein OS= Streptococcus transmembrane pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_0103 PE = 4 SV = 1 - [Q97T52_STRPN] Q97RE5 17.086 650 60.23 PTS system, fructose specific IIABC Multi- components OS =Streptococcus transmembrane pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_0877 PE = 4 SV = 1 - [Q97RE5_STRPN] Q97SV4 54.119 207 60.13 50S ribosomal protein L4 Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = rplD PE = 3 SV =1 - [RL4_STRPN] Q97RK7 19.46 12 848 59.83 Aminopeptidase N OS =Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = pepN PE = 4 SV = 1 - [Q97RK7_STRPN] Q97Q62 34.67 11424 59.21 Psr protein OS = Streptococcus Intracellular/ pneumoniaeserotype 4 (strain ATCC TMH BAA-334/TIGR4) GN = SP_1368 PE = 4 start SV= 1 - [Q97Q62_STRPN] AFTER 60 O07344 26.96 5 204 58.64 Signal peptidaseI OS = Streptococcus N- pneumoniae serotype 4 (strain ATCC terminallyBAA-334/TIGR4) GN = lepB PE = 3 SV = 2 - anchored [LEP_STRPN] (No CS)P0A3S3 34.67 6 274 57.63 DNA-entry nuclease OS = Streptococcus N-pneumoniae serotype 4 (strain ATCC terminally BAA-334/TIGR4) GN = endAPE = 1 SV = 1 - anchored [NUCE_STRPN] (No CS) Q97PU5 29.55 10 572 57.38Phosphoglucomutase Intracellular OS = Streptococcus pneumoniae serotype4 (strain ATCC BAA-334/ TIGR4) GN = pgm PE = 3 SV = 1 - [Q97PU5_STRPN]Q97RC6 31.34 8 335 57.3 ATP-dependent 6- Intracellularphosphofructokinase OS = Streptococcus pneumoniae serotype 4 (strainATCC BAA-334/ TIGR4) GN = pfkA PE = 3 SV = 1 - [PFKA_STRPN] Q97SV5 37.56 208 56.96 50S ribosomal protein L3 Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = rplC PE = 3 SV =1 - [RL3_STRPN] Q97RL9 25.56 6 399 56.21 Uncharacterized protein N- OS =Streptococcus pneumoniae terminally serotype 4 (strain ATCC BAA-334/anchored TIGR4) GN = SP_0785 PE = 4 SV = 1 - (No CS) [Q97RL9_STRPN]P66278 18.18 2 66 55.84 50S ribosomal protein L35 Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =rpmI PE = 3 SV = 1 - [RL35_STRPN] I6L8N0 40.82 12 365 55.32UDP-N-acetylglucosamine-2-epimerase Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = cps4I PE = 3 SV= 1 - [I6L8N0_STRPN] Q97PQ8 57.34 6 143 54.74 Uncharacterized proteinIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = SP_1545 PE = 4 SV = 1 - [Q97PQ8_STRPN] Q97TA6 33.6410 330 52.71 Phosphate acyltransferase Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = plsX PE = 3 SV =1 - [PLSX_STRPN] Q97PK5 27.22 2 158 52.32 Uncharacterized protein N- OS= Streptococcus pneumoniae terminally serotype 4 (strain ATCC BAA-334/anchored TIGR4) GN = SP_1604 PE = 4 SV = 1 - (No CS) [Q97PK5_STRPN]Q97SN5 67.57 8 148 51.12 50S ribosomal protein L13 Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =rplM PE = 3 SV = 1 - [Q97SN5_STRPN] Q97RV8 60.32 8 126 49.28Uncharacterized protein N- OS = Streptococcus pneumoniae terminallyserotype 4 (strain ATCC BAA-334/ anchored TIGR4) GN = SP_0678 PE = 4 SV= 1 - (No CS) [Q97RV8_STRPN] Q97R14 30.25 7 324 48.6 Uncharacterizedprotein Secretory OS = Streptococcus pneumoniae (released) serotype 4(strain ATCC BAA-334/ (with CS) TIGR4) GN = SP_1027 PE = 4 SV = 1 -[Q97R14_STRPN] P35597 15.04 7 778 48.34 Probable cation-transportingATPase Multi- exp7 OS = Streptococcus pneumoniae transmembrane serotype4 (strain ATCC BAA-334/ TIGR4) GN = exp7 PE = 3 SV = 2 - [EXP7_STRPN]P65239 41.61 9 322 48.08 Ribose-phosphate pyrophosphokinaseIntracellular 1 OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = prs1 PE = 3 SV = 1 - [KPRS1_STRPN] Q97Q37 46.13 7271 47.67 Amino acid ABC transporter, amino Lipid acid-binding proteinOS = Streptococcus anchored pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_1394 PE = 4 SV = 1 - [Q97Q37_STRPN] Q97R36 52.438 185 47.65 Thioredoxin family protein Lipid OS = Streptococcuspneumoniae anchored serotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_1000PE = 1 SV = 1 - [Q97R36_STRPN] Q97N48 26.81 5 276 45.87 Uncharacterizedprotein Intracellular/ OS = Streptococcus pneumoniae TMH serotype 4(strain ATCC BAA-334/ start TIGR4) GN = SP_2223 PE = 4 SV = 1 - AFTER 60[Q97N48_STRPN] I6L8R1 22.79 4 215 45.06 ABC transporter, ATP-bindingprotein Intracellular Vexp2 OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = vex2 PE = 3 SV = 1 - [I6L8R1_STRPN]Q97SC6 20.93 10 774 44.93 Formate acetyltransferase Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =pfl PE = 4 SV = 1 - [Q97SC6_STRPN] Q97QD5 51.08 8 186 44.75 LemA proteinOS = Streptococcus N- pneumoniae serotype 4 (strain ATCC terminallyBAA-334/TIGR4) GN = lemA PE = 1 SV = 1 - anchored [Q97QD5_STRPN] (No CS)Q97PF9 37.71 9 419 44.71 Cell division protein FtsZ Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =ftsZ PE = 3 SV = 1 - [Q97PF9_STRPN] P0A3B7 36.71 7 346 44.56 Elongationfactor Ts OS = Streptococcus Intracellular pneumoniae serotype 4 (strainATCC BAA-334/TIGR4) GN = tsf PE = 1 SV = 2 - [EFTS_STRPN] P72524 20.5610 822 44.46 DNA gyrase subunit A Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = gyrA PE = 3 SV =3 - [GYRA_STRPN] P67282 26.22 11 534 43.28 Ribonuclease Y OS =Streptococcus N- pneumoniae serotype 4 (strain ATCC terminallyBAA-334/TIGR4) GN = rny PE = 3 SV = 1 - anchored [RNY_STRPN] (No CS)P66419 38.2 6 89 42.98 30S ribosomal protein S14 Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =rpsN PE = 3 SV = 1 - [RS14_STRPN] Q97SV1 48.89 6 180 42.64 50S ribosomalprotein L5 Intracellular OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = rplE PE = 3 SV = 1 - [RL5_STRPN]Q97NW9 7.48 5 655 42.08 Trehalose PTS system, IIABC Multi- components OS= Streptococcus transmembrane pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = SP_1884 PE = 4 SV = 1 - [Q97NW9_STRPN] Q97QI8 91.066 123 40.86 Uncharacterized protein Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_1218 PE = 4SV = 1 - [Q97QI8_STRPN] Q97T04 11.92 2 193 39.59 Uncharacterized proteinMulti- OS = Streptococcus pneumoniae transmembrane serotype 4 (strainATCC BAA-334/ TIGR4) GN = SP_0157 PE = 4 SV = 1 - [Q97T04_STRPN] Q97PB89.09 4 627 39.49 PTS system IIABC components Multi- OS = Streptococcuspneumoniae transmembrane serotype 4 (strain ATCC BAA-334/ TIGR4) GN =SP_1722 PE = 4 SV = 1 - [Q97PB8_STRPN] Q97PI9 57.3 5 89 39.48 30Sribosomal protein S15 Intracellular OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = rpsO PE = 3 SV = 1 -[RS15_STRPN] P61182 40.35 4 114 39.35 50S ribosomal protein L22Intracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = rplV PE = 1 SV = 1 - [RL22_STRPN] Q97SU7 51.69 6178 38.14 50S ribosomal protein L6 Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = rplF PE = 3 SV =1 - [RL6_STRPN] Q97NB9 39.1 8 335 38.04 Uncharacterized protein Multi-OS = Streptococcus pneumoniae transmembrane serotype 4 (strain ATCCBAA-334/ TIGR4) GN = SP_2132 PE = 4 SV = 1 - [Q97NB9_STRPN] Q97QP7 5.348 2004 37.84 Immunoglobulin A1 protease Multi- OS = Streptococcuspneumoniae transmembrane serotype 4 (strain ATCC BAA-334/ TIGR4) GN =iga PE = 3 SV = 1 - [IGA1A_STRPN] Q97N55 23.72 6 392 37.16 Secreted 45kd protein Secretory OS = Streptococcus pneumoniae (released) serotype 4(strain ATCC BAA-334/ (with CS) TIGR4) GN = usp45 PE = 4 SV = 1 -[Q97N55_STRPN] P66942 18.25 4 252 36.63 Triosephosphate isomeraseIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = tpiA PE = 3 SV = 1 - [TPIS_STRPN] Q97QC6 66.09 7115 35.81 50S ribosomal protein L19 Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = rplS PE = 3 SV =1 - [RL19_STRPN] I6L8S7 14.6 5 459 35.54 Transmembrane protein Vexp3Multi- OS = Streptococcus pneumoniae transmembrane serotype 4 (strainATCC BAA-334/ TIGR4) GN = vex3 PE = 4 SV = 1 - [I6L8S7_STRPN] P3559512.4 6 726 35.44 PTS system glucose-specific EIICBA Multi- component OS= Streptococcus transmembrane pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = exp5 PE = 3 SV = 2 - [PTG3C_STRPN] Q97PV1 14.88 3289 34.89 Putative glycerol uptake facilitator Multi- protein OS =Streptococcus pneumoniae transmembrane serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1491 PE = 3 SV = 1 - [Q97PV1_STRPN] Q97NM6 21.04 6 40433.79 Aminotransferase, class I Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_1994 PE = 4SV = 1 - [Q97NM6_STRPN] Q97SD2 45.93 5 246 33.44 Amino acid ABCtransporter, ATP- Intracellular binding protein OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_0452 PE = 3 SV= 1 - [Q97SD2_STRPN] Q97NN3 36.15 6 213 33.1 ABC transporter,ATP-binding protein Intracellular OS = Streptococcus pneumoniae serotype4 (strain ATCC BAA-334/ TIGR4) GN = SP_1987 PE = 1 SV = 1 -[Q97NN3_STRPN] Q97RJ0 23.77 5 244 33.09 Amino acid ABC transporter, ATP-Intracellular binding protein OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/TIGR4) GN = SP_0824 PE = 3 SV = 1 - [Q97RJ0_STRPN]Q97NJ7 27.27 3 99 32.76 Preprotein translocase, YajC subunit N- OS =Streptococcus pneumoniae terminally serotype 4 (strain ATCC BAA-334/anchored TIGR4) GN = yajC-2 PE = 4 SV = 1 - (No CS) [Q97NJ7_STRPN]P66095 34.5 8 229 32.4 50S ribosomal protein L1 Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =rplA PE = 3 SV = 1 - [RL1_STRPN] Q97NT7 38.61 6 158 32.3 Uncharacterizedprotein Multi- OS = Streptococcus pneumoniae transmembrane serotype 4(strain ATCC BAA-334/ TIGR4) GN = SP_1926 PE = 4 SV = 1 - [Q97NT7_STRPN]P63667 29.5 4 139 31.7 ATP synthase epsilon chain Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =atpC PE = 3 SV = 1 - [ATPE_STRPN] Q97T58 18.08 4 354 31.6 Conserveddomain protein Multi- OS = Streptococcus pneumoniae transmembraneserotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_0097 PE = 4 SV = 1 -[Q97T58_STRPN] Q97R06 36.74 6 264 31.17 Iron-compound ABC transporter,ATP- Intracellular binding protein OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1035 PE = 3 SV = 1 -[Q97R06_STRPN] Q97PQ5 20.34 6 531 30.81 Uncharacterized protein Multi-OS = Streptococcus pneumoniae transmembrane serotype 4 (strain ATCCBAA-334/ TIGR4) GN = SP_1548 PE = 4 SV = 1 - [Q97PQ5_STRPN] Q97R44 33.912 115 30.62 Uncharacterized protein Intracellular/ OS = Streptococcuspneumoniae TMH serotype 4 (strain ATCC BAA-334/ start TIGR4) GN =SP_0990 PE = 4 SV = 1 - AFTER 60 [Q97R44_STRPN] Q97SW3 47.73 3 88 29.98Uncharacterized protein Intracellular OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_0201 PE = 4 SV = 1 -[Q97SW3_STRPN] I6L8N6 27.01 4 211 29.5 Capsular polysaccharidebiosynthesis N- protein Cps4E OS = Streptococcus terminally pneumoniaeserotype 4 (strain ATCC anchored BAA-334/TIGR4) GN = cps4E PE = 4 SV =1 - (No CS) [I6L8N6_STRPN] Q97SU3 34.25 5 146 28.53 50S ribosomalprotein L15 Intracellular OS = Streptococcus pneumoniae serotype 4(strain ATCC BAA-334/ TIGR4) GN = rplO PE = 3 SV = 1 - [RL15_STRPN]Q97QS6 25.86 6 379 28.15 Glycogen biosynthesis protein GlgDIntracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = glgD PE = 4 SV = 1 - [Q97QS6_STRPN] Q97P39 52.24 267 27.23 UPF0337 protein SP_1805 Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_1805 PE = 3SV = 1 - [Y1805_STRPN] P06653 20.44 6 318 25.81 Autolysin OS =Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = lytA PE = 1 SV = 2 - [ALYS_STRPN] Q97PX9 12.78 5 26625.41 Amino acid ABC transporter, permease Multi- protein OS =Streptococcus pneumoniae transmembrane serotype 4 (strain ATCC BAA-334/TIGR4) GN = SP_1461 PE = 3 SV = 1 - [Q97PX9_STRPN] Q97QW4 17.65 5 42524.84 Uncharacterized protein Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_1083 PE = 4SV = 1 - [Q97QW4_STRPN] Q97S93 13.27 7 535 23.79 CTP synthase OS =Streptococcus Intracellular pneumoniae serotype 4 (strain ATCCBAA-334/TIGR4) GN = pyrG PE = 3 SV = 1 - [PYRG_STRPN] Q97SX1 26.14 2 8822 UPF0297 protein SP_0192 Intracellular OS = Streptococcus pneumoniaeserotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_0192 PE = 1 SV = 1 -[Y192_STRPN] Q97RH7 50 4 78 21.34 30S ribosomal protein S20Intracellular OS = Streptococcus pneumoniae serotype 4 (strain ATCCBAA-334/ TIGR4) GN = rpsT PE = 3 SV = 1 - [RS20_STRPN] P66050 45.78 5166 20.98 50S ribosomal protein L10 Intracellular OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = rplJ PE = 3 SV =2 - [RL10_STRPN] Q97RW6 32.14 3 56 20.85 Uncharacterized proteinSecretory OS = Streptococcus pneumoniae (released) serotype 4 (strainATCC BAA-334/ (with CS) TIGR4) GN = SP_0670 PE = 4 SV = 1 -[Q97RW6_STRPN] Q9FBB7 29.41 4 255 20.65 Acetyl-coenzyme A carboxylaseIntracellular carboxyl transferase subunit alpha OS = Streptococcuspneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN = accA PE = 1 SV =1 - [ACCA_STRPN] I6L8V8 30.45 4 243 20.58 3-oxoacyl-[acyl-carrierprotein] Intracellular reductase OS = Streptococcus pneumoniae serotype4 (strain ATCC BAA-334/TIGR4) GN = fabG PE = 3 SV = 1 - [I6L8V8_STRPN]P66581 41.46 4 164 20.46 30S ribosomal protein S5 Intracellular OS =Streptococcus pneumoniae serotype 4 (strain ATCC BAA-334/ TIGR4) GN =rpsE PE = 3 SV = 1 - [RS5_STRPN] Q97T45 11.82 5 694 20.39Uncharacterized protein Multi- OS = Streptococcus pneumoniaetransmembrane serotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_0110 PE =4 SV = 1 - [Q97T45_STRPN] Q97P18 18.31 4 355 20.06 ABC transporter,substrate-binding Lipid protein OS = Streptococcus pneumoniae anchoredserotype 4 (strain ATCC BAA-334/ TIGR4) GN = SP_1826 PE = 4 SV = 1 -[Q97P18_STRPN]

TABLE 5 Proteins identified in MV_(L) from serotype 3. The accessionnumbers refer to SwissProt on the date of filing. Accession # Coverage #Peptides #AAs Score Description Localization B2IQV2 91.09 50 404 5251.94Elongation factor Tu Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = tuf PE = 3 SV = 1 - [B2IQV2_STRPS] B2IPX8 95.62 40 4342291.84 Enolase OS = Streptococcus Intracellular pneumoniae (strainCGSP14) GN = eno PE = 3 SV = 1 - [ENO_STRPS] B2INW2 70.86 42 374 2221.7Lipoprotein OS = Streptococcus Lipid pneumoniae (strain CGSP14) anchoredGN = SPCG_0787 PE = 4 SV = 1 - [B2INW2_STRPS] B2ISJ9 85.28 67 6931851.66 Elongation factor G Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = fusA PE = 3 SV = 1 - [EFG_STRPS] B2IMI7 73.82 33359 1675.28 Glyceraldehyde-3-phosphate Intracellular dehydrogenase OS =Streptococcus pneumoniae (strain CGSP14) GN = gapA PE = 3 SV = 1 -[B2IMI7_STRPS] B2IM39 82.94 101 1225 1348.46 DNA-directed RNA polymeraseIntracellular subunit beta′ OS = Streptococcus pneumoniae (strainCGSP14) GN = rpoC PE = 3 SV = 1 - [RPOC_STRPS] B2IMJ9 86.85 68 8901321.16 Aldehyde-alcohol dehydrogenase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = adhE PE = 3 SV = 1 - [B2IMJ9_STRPS]B2IPD4 73.16 38 313 1171.98 Foldase protein PrsA Lipid OS =Streptococcus pneumoniae anchored (strain CGSP14) GN = prsA PE = 3 SV =1 - [PRSA_STRPS] B2INY1 91 53 400 1141.84 30S ribosomal protein S1Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rpsA PE= 4 SV = 1 - [B2INY1_STRPS] B2IP53 87.46 27 335 830.14 ATP-dependent 6-Intracellular phosphofructokinase OS = Streptococcus pneumoniae (strainCGSP14) GN = pfkA PE = 3 SV = 1 - [PFKA_STRPS] B2IQX0 90.6 38 468 772.66ATP synthase subunit beta Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = atpD PE = 3 SV = 1 - [ATPB_STRPS] B2IRV7 72.76 62837 760.12 Protein translocase subunit SecA Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = secA PE = 3 SV = 1 -[B2IRV7_STRPS] B2IN25 78.22 51 450 751.01 Maltose/maltodextrin ABC Lipidtransporter, maltose/maltodextrin- anchored binding protein OS =Streptococcus pneumoniae (strain CGSP14) GN = SPCG_2072 PE = 4 SV = 1 -[B2IN25_STRPS] B2INB2 72.76 41 591 749.62 Pyruvate oxidase OS =Streptococcus Intracellular pneumoniae (strain CGSP14) GN = spxB PE = 3SV = 1 - [B2INB2_STRPS] B2ISZ0 69.04 47 659 746.61 Oligopeptide ABCtransporter, Lipid oligopeptide-binding protein AmiA anchored OS =Streptococcus pneumoniae (strain CGSP14) GN = amiA PE = 4 SV = 1 -[B2ISZ0_STRPS] B2ISK8 56.44 22 303 734.11 PTS system, mannose-specificIID Multi- component OS = Streptococcus transmembrane pneumoniae (strainCGSP14) GN = manN PE = 4 SV = 1 - [B2ISK8_STRPS] B2INN2 85.33 30 259682.23 30S ribosomal protein S2 Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = rpsB PE = 3 SV = 2 - [RS2_STRPS] B2IP5476.05 39 501 676.55 Pyruvate kinase OS = Streptococcus Intracellularpneumoniae (strain CGSP14) GN = SPCG_0873 PE = 3 SV = 1 - [B2IP54_STRPS]B2IR47 69.48 50 652 673.91 Oligopeptide ABC transporter, Lipidoligopeptide-binding protein AliB anchored OS = Streptococcus pneumoniae(strain CGSP14) GN = aliB PE = 4 SV = 1 - [B2IR47_STRPS] B2IR00 66.26 44652 636.95 ATP-dependent zinc Multi- metalloprotease FtsH transmembraneOS = Streptococcus pneumoniae (strain CGSP14) GN = ftsH PE = 3 SV = 1 -[B2IR00_STRPS] B2IM40 77.67 50 824 633.52 DNA-directed RNA polymeraseIntracellular subunit beta OS = Streptococcus pneumoniae (strain CGSP14)GN = rpoB PE = 3 SV = 1 - [B2IM40_STRPS] B2IS43 70.76 23 277 575.93 50Sribosomal protein L2 Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = rplB PE = 3 SV = 1 - [RL2_STRPS] B2IS46 79.26 27 217 569.5630S ribosomal protein S3 Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = rpsC PE = 3 SV = 1 - [RS3_STRPS] B2IME4 64.95 51930 553.42 Translation initiation factor IF-2 Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = infB PE = 3 SV = 1 -[IF2_STRPS] B2IQY1 73.14 43 551 550.86 Uncharacterized proteinIntracellular/ OS = Streptococcus pneumoniae TMH (strain CGSP14) GN =SPCG_1503 start PE = 4 SV = 1 - [B2IQY1_STRPS] AFTER 60 B2INT1 58.9 34584 531.45 Septation ring formation regulator N- EzrA OS = Streptococcusterminally pneumoniae (strain CGSP14) anchored GN = ezrA PE = 3 SV = 1 -(No CS) [B2INT1_STRPS] B2IPY8 67.93 32 474 511.28Glyceraldehyde-3-phosphate Intracellular dehydrogenase, NADP-dependentOS = Streptococcus pneumoniae (strain CGSP14) GN = gapN PE = 3 SV = 1 -[B2IPY8_STRPS] B2IRQ6 79.23 20 313 461.56 Manganese ABC transporter,Lipid manganese-binding adhesion anchored liprotein OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_1623 PE = 3 SV = 1 - [B2IRQ6_STRPS]B2IPU8 63.14 36 719 457.32 Ribonucleoside-diphosphate Intracellularreductase OS = Streptococcus pneumoniae (strain CGSP14) GN = nrdE PE = 3SV = 1 - [B2IPU8_STRPS] B2IM49 72.42 27 330 449.87 Aspartate--ammonialigase Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =asnA PE = 3 SV = 1 - [ASNA_STRPS] B2IQX2 68.86 31 501 447.54 ATPsynthase subunit alpha Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = atpA PE = 3 SV = 1 - [ATPA_STRPS] B2IMK3 70.36 32658 435.4 Transketolase OS = Streptococcus Intracellular pneumoniae(strain CGSP14) GN = tktA PE = 3 SV = 1 - [B2IMK3_STRPS] B2IRH9 64.45 36616 434.01 Capsular polysaccharide Multi- biosynthesis protein, putativetransmembrane OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_0098 PE = 4 SV = 1 - [B2IRH9_STRPS] B2IN95 82.06 37 496 432.16Lysine--tRNA ligase Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = lysS PE = 3 SV = 1 - [SYK_STRPS] B2IS90 71.32 40 537 413.57Ribonuclease Y OS = Streptococcus Intracellular pneumoniae (strainCGSP14) GN = rny PE = 3 SV = 1 - [B2IS90_STRPS] B2IRT9 66.06 32 442412.96 Sugar ABC transporter, sugar- Lipid binding protein OS =Streptococcus anchored pneumoniae (strain CGSP14) GN = SPCG_1656 PE = 4SV = 1 - [B2IRT9_STRPS] B2IN10 73.37 15 338 401 Glycerol-3-phosphateIntracellular dehydrogenase [NAD(P)+] OS = Streptococcus pneumoniae(strain CGSP14) GN = gpsA PE = 3 SV = 1 - [GPDA_STRPS] B2IP07 38.46 23650 398.03 PTS system, fructose specific IIABC Multi- components OS =Streptococcus transmembrane pneumoniae (strain CGSP14) GN = fruA PE = 4SV = 1 - [B2IP07_STRPS] B2IPB5 52.94 10 119 388.03 50S ribosomal proteinL20 Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =rplT PE = 3 SV = 1 - [RL20_STRPS] B2ISI9 66.29 35 617 385.71Proline--tRNA ligase Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = proS PE = 3 SV = 1 - [SYP_STRPS] B2INH5 83.95 31 299 370.13SPFH domain/Band 7 family N- OS = Streptococcus pneumoniae terminally(strain CGSP14) GN = SPCG_2124 anchored PE = 4 SV = 1 - [B2INH5_STRPS](No CS) B2IQ40 64.14 46 898 367.23 Phosphoenolpyruvate carboxylaseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = ppc PE= 3 SV = 1 - [B2IQ40_STRPS] B2IML8 76.35 25 406 358.33 Acetate kinase OS= Streptococcus Intracellular pneumoniae (strain CGSP14) GN = ackA PE =3 SV = 1 - [B2IML8_STRPS] B2IRS1 53.94 28 419 341.4 Cell divisionprotein FtsZ Intracellular OS = Streptococcus pneumoniae (strain CGSP14)GN = ftsZ PE = 3 SV = 1 - [B2IRS1_STRPS] B2IMN0 96.59 22 352 339.61Alcohol dehydrogenase, zinc- Intracellular containing OS = Streptococcuspneumoniae (strain CGSP14) GN = adh PE = 3 SV = 1 - [B2IMN0_STRPS]B2ISJ7 51.09 13 137 337.74 30S ribosomal protein S12 Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = rpsL PE = 3 SV = 1 -[RS12_STRPS] B2IS41 63.29 16 207 337.71 50S ribosomal protein L4Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rplD PE= 3 SV = 1 - [RL4_STRPS] B2ILW7 80.68 18 414 332.023-oxoacyl-[acyl-carrier-protein] Intracellular synthase 2 OS =Streptococcus pneumoniae (strain CGSP14) GN = fabF PE = 3 SV = 1 -[B2ILW7_STRPS] B2IRM9 70.29 22 276 330.56 ABC transporter,substrate-binding Lipid protein OS = Streptococcus anchored pneumoniae(strain CGSP14) GN = SPCG_0152 PE = 4 SV = 1 - [B2IRM9_STRPS] B2ISY947.99 21 498 319.21 Oligopeptide ABC transporter, Multi- permeaseprotein AmiC transmembrane OS = Streptococcus pneumoniae (strain CGSP14)GN = amiC PE = 3 SV = 1 - [B2ISY9_STRPS] B2IQT6 62.75 22 459 313.52 NADHoxidase OS = Streptococcus Intracellular pneumoniae (strain CGSP14) GN =nox PE = 4 SV = 1 - [B2IQT6_STRPS] B2IPP2 47.02 32 721 312 Amino acidABC transporter, amino Multi- acid-binding protein/permeasetransmembrane protein OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1062 PE = 3 SV = 1 - [B2IPP2_STRPS] B2IS47 63.5 13 137 300.82 50Sribosomal protein L16 Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = rplP PE = 3 SV = 1 - [RL16_STRPS] B2IRC1 66.67 19339 299.62 UDP-glucose 4-epimerase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = galE PE = 4 SV = 1 - [B2IRC1_STRPS]B2IQW3 71.22 25 278 296.15 Amino acid ABC transporter, amino Lipidacid-binding protein anchored OS = Streptococcus pneumoniae (strainCGSP14) GN = SPCG_1485 PE = 4 SV = 1 - [B2IQW3_STRPS] B2IRR1 67.83 19230 295.06 2,3-bisphosphoglycerate- Intracellular dependentphosphoglycerate mutase OS = Streptococcus pneumoniae (strain CGSP14) GN= gpmA PE = 3 SV = 1 - [GPMA_STRPS] B2IS34 61.87 39 737 290.83 Anaerobicribonucleoside Intracellular triphosphate reductase OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_0212 PE = 4 SV = 1 - [B2IS34_STRPS]B2IS52 87.78 18 180 288.42 50S ribosomal protein L5 Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = rplE PE = 3 SV = 1 -[RL5_STRPS] B2IQU1 65.63 33 678 288.36 Glycine--tRNA ligase beta subunitIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = glyS PE= 3 SV = 1 - [SYGB_STRPS] B2ILY5 60.36 21 555 284.08 Uncharacterizedprotein Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_0438 PE = 4 SV = 1 - [B2ILY5_STRPS] B2IS40 59.62 14 208 283.69 50Sribosomal protein L3 Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = rplC PE = 3 SV = 1 - [RL3_STRPS] B2IM73 32.44 18 521 281.46Amino acid ABC transporter, amino Multi- acid-binding protein/permeasetransmembrane protein OS = Streptococcus pneumoniae (strain CGSP14) GN =glnH PE = 3 SV = 1 - [B2IM73_STRPS] B2IRI9 52.65 17 302 275.76 Aminoacid ABC transporter, Lipid periplasmic amino acid-binding anchoredprotein, putative OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_0108 PE = 4 SV = 1 - [B2IRI9_STRPS] B2IRN0 57.75 22 284 275.2Lipoprotein OS = Streptococcus Lipid pneumoniae (strain CGSP14) anchoredGN = SPCG_0153 PE = 3 SV = 1 - [B2IRN0_STRPS] B2IM81 70.93 36 774 270.81Formate acetyltransferase Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = pfl PE = 4 SV = 1 - [B2IM81_STRPS] B2IRN7 65.61 30660 269.02 Threonine--tRNA ligase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = thrS PE = 3 SV = 1 - [B2IRN7_STRPS]B2IMN6 63.36 22 494 268.42 Threonine synthase Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = thrC PE = 4 SV = 1 -[B2IMN6_STRPS] B2IR98 69.68 20 376 261.66 Sugar ABC transporter,ATP-binding Intracellular protein OS = Streptococcus pneumoniae (strainCGSP14) GN = SPCG_1565 PE = 3 SV = 1 - [B2IR98_STRPS] B2IMG8 58.42 23404 253.09 Aspartate aminotransferase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = aspC PE = 4 SV = 1 - [B2IMG8_STRPS]B2ILW6 65.43 13 243 252.82 3-ketoacyl-(Acyl-carrier-protein)Intracellular reductase OS = Streptococcus pneumoniae (strain CGSP14) GN= fabG PE = 4 SV = 1 - [B2ILW6_STRPS] B2INF2 70.18 25 513 248.48 ABCtransporter, ATP-binding Intracellular protein OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_0719 PE = 4 SV = 1 - [B2INF2_STRPS]B2IRS2 78.12 20 457 247.37 Cell division protein FtsA Multi- OS =Streptococcus pneumoniae transmembrane (strain CGSP14) GN = ftsZ PE = 3SV = 1 - [B2IRS2_STRPS] B2IM41 70.82 22 377 246.54 Uncharacterizedprotein Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1926 PE = 3 SV = 1 - [B2IM41_STRPS] B2IQ88 66.09 13 115 240.71 50Sribosomal protein L19 Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = rplS PE = 3 SV = 1 - [RL19_STRPS] B2IQI0 56.1 22344 240.32 Uncharacterized protein Multi- OS = Streptococcus pneumoniaetransmembrane (strain CGSP14) GN = SPCG_1352 PE = 4 SV = 1 -[B2IQI0_STRPS] B2IND2 63.73 18 386 236.96 Branched-chain amino acid ABCLipid transporter, amino acid-binding anchored protein OS =Streptococcus pneumoniae (strain CGSP14) GN = SPCG_0699 PE = 4 SV = 1 -[B2IND2_STRPS] B2INP5 71.34 24 492 236.94 Inosine-5′-monophosphateIntracellular dehydrogenase OS = Streptococcus pneumoniae (strainCGSP14) GN = imdH PE = 3 SV = 1 - [B2INP5_STRPS] B2ISL2 67.26 18 339235.53 Alcohol dehydrogenase Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = adhP PE = 3 SV = 1 - [B2ISL2_STRPS] B2ISM4 59.23 9130 228.51 30S ribosomal protein S9 Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = rpsI PE = 3 SV = 1 - [RS9_STRPS] B2IR7140.26 23 914 227.01 Cation-transporting ATPase, E1-E2 Multi- family OS =Streptococcus transmembrane pneumoniae (strain CGSP14) GN = pacL PE = 3SV = 1 - [B2IR71_STRPS] B2IPP1 85.77 17 246 225.41 Amino acid ABCtransporter, ATP- Intracellular binding protein OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_1061 PE = 4 SV = 1 - [B2IPP1_STRPS]B2ISY7 58.31 18 355 221.22 Oligopeptide ABC transporter, ATP-Intracellular binding protein AmiE OS = Streptococcus pneumoniae (strainCGSP14) GN = amiE PE = 3 SV = 1 - [B2ISY7_STRPS] B2IQL1 56.83 16 271219.73 Amino acid ABC transporter, amino Lipid acid-binding proteinanchored OS = Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1383 PE= 4 SV = 1 - [B2IQL1_STRPS] B2ISG3 77.23 19 202 219.45 General stressprotein 24, putative Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = SPCG_1781 PE = 4 SV = 1 - [B2ISG3_STRPS] B2IMA2 70.1 20 398216.35 Phosphoglycerate kinase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = pgk PE = 3 SV = 1 - [PGK_STRPS] B2IN7149.03 25 620 212.01 Elongation factor Tu family protein Intracellular OS= Streptococcus pneumoniae (strain CGSP14) GN = SPCG_0638 PE = 4 SV =1 - [B2IN71_STRPS] B2IPR1 75.6 16 332 210.14 L-lactate dehydrogenaseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = Idh PE= 3 SV = 1 - [B2IPR1_STRPS] B2IQK0 51.83 30 872 209.16 Alanine--tRNAligase Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =alaS PE = 3 SV = 1 - [SYA_STRPS] B2IS44 61.29 12 93 208.67 30S ribosomalprotein S19 Intracellular OS = Streptococcus pneumoniae (strain CGSP14)GN = rpsS PE = 3 SV = 1 - [RS19_STRPS] B2IMN9 48.47 17 425 208.24Uncharacterized protein Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = SPCG_2034 PE = 4 SV = 1 - [B2IMN9_STRPS] B2IR1772.27 16 339 208.13 Ribose-phosphate Intracellular pyrophosphokinase OS= Streptococcus pneumoniae (strain CGSP14) GN = prsA PE = 3 SV = 1 -[B2IR17_STRPS] B2IQR1 62.69 25 520 208 GMP synthase [glutamine-Intracellular hydrolyzing] OS = Streptococcus pneumoniae (strain CGSP14)GN = guaA PE = 3 SV = 1 - [GUAA_STRPS] B2INL3 50.25 23 810 202.66ATP-dependent Clp protease, ATP- Intracellular binding subunit OS =Streptococcus pneumoniae (strain CGSP14) GN = SPCG_2162 PE = 3 SV = 1 -[B2INL3_STRPS] B2IN11 58.53 17 299 197.31 UTP--glucose-1-phosphateSecretory uridylyltransferase (released) OS = Streptococcus pneumoniae(with (strain CGSP14) GN = galU PE = 3 SV = 1 - CS) [B2IN11_STRPS]B2IQU5 44.38 17 480 197.17 Peptidoglycan N-acetylglucosamine N-deacetylase A OS = Streptococcus terminally pneumoniae (strain CGSP14)anchored GN = pgdA PE = 4 SV = 1 - (No CS) [B2IQU5_STRPS] B2INE4 57.4919 447 196.45 DEAD-box ATP-dependent RNA Intracellular helicase CshB OS= Streptococcus pneumoniae (strain CGSP14) GN = rheB PE = 3 SV = 1 -[B2INE4_STRPS] B2IMU6 65.97 15 238 196.17 Uncharacterized protein LipidOS = Streptococcus pneumoniae anchored (strain CGSP14) GN = SPCG_0590 PE= 4 SV = 1 - [B2IMU6_STRPS] B2INT7 54.92 27 752 195.47 ATP-dependent Clpprotease, ATP- Intracellular binding subunit ClpE OS = Streptococcuspneumoniae (strain CGSP14) GN = clpE PE = 3 SV = 1 - [B2INT7_STRPS]B2IM28 44.59 17 388 194.98 Protein RecA OS = Streptococcus Intracellularpneumoniae (strain CGSP14) GN = recA PE = 3 SV = 1 - [RECA_STRPS] B2IQL479.78 18 267 192.86 Phosphate import ATP-binding Intracellular proteinPstB OS = Streptococcus pneumoniae (strain CGSP14) GN = pstB PE = 3 SV =1 - [B2IQL4_STRPS] B2ISL0 43.67 13 332 190.43 PTS system,mannose-specific IIAB Intracellular components OS = Streptococcuspneumoniae (strain CGSP14) GN = manL PE = 4 SV = 1 - [B2ISL0_STRPS]B2INW3 54.6 19 511 188.38 Sugar ABC transporter, ATP-bindingIntracellular protein OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_0788 PE = 4 SV = 1 - [B2INW3_STRPS] B2INT0 40.9 22 648 185.21 DNAgyrase subunit B Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = gyrB PE = 3 SV = 1 - [B2INT0_STRPS] B2IPI7 57.18 13 341184.52 Iron-compound ABC transporter, Lipid iron compound-bindingprotein anchored OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1009 PE = 4 SV = 1 - [B2IPI7_STRPS] B2IPT0 48.09 22 628 183.98Elongation factor 4 Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = lepA PE = 3 SV = 1 - [B2IPT0_STRPS] B2IN23 39.89 17 752181.97 Alpha-1,4 glucan phosphorylase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = malP PE = 3 SV = 1 - [B2IN23_STRPS]B2IQZ2 64.71 20 374 180.03 Ribosome-binding ATPase YchF Intracellular OS= Streptococcus pneumoniae (strain CGSP14) GN = ychF PE = 3 SV = 1 -[B2IQZ2_STRPS] B2IMU8 66.81 17 229 179.85 50S ribosomal protein L1Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rplA PE= 3 SV = 1 - [RL1_STRPS] B2ISY6 51.06 14 331 178.98 Oligopeptide ABCtransporter, ATP- Intracellular binding protein AmiF OS = Streptococcuspneumoniae (strain CGSP14) GN = amiF PE = 3 SV = 1 - [B2ISY6_STRPS]B2IR87 54.15 18 410 170.5 ATP-dependent Clp protease ATP- Intracellularbinding subunit ClpX OS = Streptococcus pneumoniae (strain CGSP14) GN =clpX PE = 3 SV = 1 - [CLPX_STRPS] B2IR07 58.6 24 442 169.98Adenylosuccinate synthetase Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = purA PE = 3 SV = 1 - [B2IR07_STRPS] B2ILV9 61.23 20454 169.93 Aspartokinase OS = Streptococcus Intracellular pneumoniae(strain CGSP14) GN = lysC PE = 3 SV = 1 - [B2ILV9_STRPS] B2IMZ6 42.12 25857 169.84 DNA mismatch repair protein MutS Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = hexA PE = 3 SV = 1 -[B2IMZ6_STRPS] B2IS57 84.76 17 164 169.34 30S ribosomal protein S5Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rpsE PE= 3 SV = 1 - [B2IS57_STRPS] B2ILR1 29.62 20 719 166.63Penicillin-binding protein 1A N- OS = Streptococcus pneumoniaeterminally (strain CGSP14) GN = pbp1A PE = 4 anchored SV = 1 -[B2ILR1_STRPS] (No CS) B2IQ78 53.73 22 523 165.23 Signal recognitionparticle protein Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = ffh PE = 3 SV = 1 - [B2IQ78_STRPS] B2ISM3 78.38 17 148163.82 50S ribosomal protein L13 Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = rplM PE = 3 SV = 1 - [B2ISM3_STRPS]B2INJ2 70.02 21 427 163.61 DltD protein OS = Streptococcus N- pneumoniae(strain CGSP14) terminally GN = dltD PE = 4 SV = 1 - anchored[B2INJ2_STRPS] (No CS) B2ISJ1 48.84 19 602 159.21Glutamine--fructose-6-phosphate Intracellular aminotransferase[isomerizing] OS = Streptococcus pneumoniae (strain CGSP14) GN = glmS PE= 3 SV = 1 - [B2ISJ1_STRPS] B2IPM7 52.69 18 577 159.04Phosphoenolpyruvate-protein Intracellular phosphotransferase OS =Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1121 PE = 3 SV = 1 -[B2IPM7_STRPS] B2IQ29 61.01 17 436 157.98 GTPase Obg OS = StreptococcusIntracellular pneumoniae (strain CGSP14) GN = obg PE = 3 SV = 1 -[OBG_STRPS] B2IS66 74.8 9 127 157.55 30S ribosomal protein S11Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rpsK PE= 3 SV = 1 - [RS11_STRPS] B2IQ67 33.36 24 1058 157.35Carbamoyl-phosphate synthase Intracellular large chain OS =Streptococcus pneumoniae (strain CGSP14) GN = carB PE = 3 SV = 1 -[CARB_STRPS] B2IQ25 62.29 16 419 154.75 UDP-N-acetylglucosamine 1-Intracellular carboxyvinyltransferase OS = Streptococcus pneumoniae(strain CGSP14) GN = murZ PE = 3 SV = 1 - [B2IQ25_STRPS] B2IQX4 82.93 14164 154.18 ATP synthase subunit b N- OS = Streptococcus pneumoniaeterminally (strain CGSP14) GN = atpF PE = 3 SV = 1 - anchored[ATPF_STRPS] (No CS) B2IM72 69.51 10 246 154.11 Amino acid ABCtransporter, ATP- Intracellular binding protein OS = Streptococcuspneumoniae (strain CGSP14) GN = glnQ PE = 4 SV = 1 - [B2IM72_STRPS]B2INR2 35.79 19 679 153.3 Methionine--tRNA ligase Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = metG PE = 3 SV = 1 -[B2INR2_STRPS] B2IS42 81.63 6 98 152.99 50S ribosomal protein L23Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rplW PE= 3 SV = 1 - [RL23_STRPS] B2IPI0 44.02 18 418 152.53 Serinehydroxymethyltransferase Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = glyA PE = 3 SV = 1 - [GLYA_STRPS] B2IRG4 73.89 21203 151.28 30S ribosomal protein S4 Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = rpsD PE = 3 SV = 1 - [RS4_STRPS] B2ILV759.29 21 452 150.33 Serine--tRNA ligase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = serS PE = 3 SV = 1 - [B2ILV7_STRPS]B2INC8 60.65 13 216 146.88 Uracil phosphoribosyltransferaseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = upp PE= 3 SV = 1 - [B2INC8_STRPS] B2IQL3 70.63 16 252 145.61 Phosphate importATP-binding Intracellular protein PstB OS = Streptococcus pneumoniae(strain CGSP14) GN = pstB PE = 3 SV = 1 - [B2IQL3_STRPS] B2IMZ5 31.91 19564 144.21 ABC transporter, ATP- Multi- binding/permease proteintransmembrane OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_2042 PE = 4 SV = 1 - [B2IMZ5_STRPS] B2IS39 59.8 8 102 143.18 30Sribosomal protein S10 Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = rpsJ PE = 3 SV = 1 - [RS10_STRPS] B2IP78 53.87 13375 141.59 Carboxynorspermidine Intracellular decarboxylase OS =Streptococcus pneumoniae (strain CGSP14) GN = nspC PE = 4 SV = 1 -[B2IP78_STRPS] B2IM60 54.81 13 416 139.41 Diaminopimelate decarboxylaseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = lysA PE= 3 SV = 1 - [B2IM60_STRPS] B2IMH3 65.03 14 346 139.16 Catabolitecontrol protein A Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = ccpA PE = 4 SV = 1 - [B2IMH3_STRPS] B2IS68 57.03 8 128139.07 50S ribosomal protein L17 Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = rplQ PE = 3 SV = 1 - [RL17_STRPS] B2INE034.95 13 329 136.77 Cell division protein FtsX Multi- OS = Streptococcuspneumoniae transmembrane (strain CGSP14) GN = ftsX PE = 3 SV = 1 -[B2INE0_STRPS] B2ILV2 41.65 20 778 134.89 Endonuclease MutS2Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = mutS2PE = 3 SV = 1 - [MUTS2_STRPS] B2IQU4 63.93 15 280 133.96 Oxidoreductase,aldo/keto Intracellular reductase family OS = Streptococcus pneumoniae(strain CGSP14) GN = SPCG_1466 PE = 4 SV = 1 - [B2IQU4_STRPS] B2IS5571.35 14 178 129.78 50S ribosomal protein L6 Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = rplF PE = 3 SV = 1 -[RL6_STRPS] B2ISJ8 67.95 15 156 129.16 30S ribosomal protein S7Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rpsG PE= 3 SV = 1 - [RS7_STRPS] B2IRR6 24.81 8 266 128.96 Cell division proteinDivIVA Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1633 PE = 4 SV = 1 - [B2IRR6_STRPS] B2ISI8 43.68 14 419 128.49 Zincmetalloprotease Multi- OS = Streptococcus pneumoniae transmembrane(strain CGSP14) GN = eep PE = 3 SV = 1 - [B2ISI8_STRPS] B2INF3 47.94 9267 128.19 Peptidyl-prolyl cis-trans isomerase Lipid OS = Streptococcuspneumoniae anchored (strain CGSP14) GN = ppiA PE = 3 SV = 1 -[B2INF3_STRPS] B2IR96 96.59 13 176 127.15 Adeninephosphoribosyltransferase Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = apt PE = 3 SV = 1 - [B2IR96_STRPS] B2IQX1 63.36 12292 124.75 ATP synthase gamma chain Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = atpG PE = 3 SV = 1 - [ATPG_STRPS] B2IQL740.41 12 292 124.49 Phosphate ABC transporter, Lipid phosphate-bindingprotein, anchored putative OS = Streptococcus pneumoniae (strain CGSP14)GN = SPCG_1389 PE = 4 SV = 1 - [B2IQL7_STRPS] B2IS49 69.77 10 86 124.4730S ribosomal protein S17 Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = rpsQ PE = 3 SV = 1 - [RS17_STRPS] B2IQV3 35.99 8289 123.64 Glycerol uptake facilitator protein, Multi- putative OS =Streptococcus transmembrane pneumoniae (strain CGSP14) GN = glpF PE = 3SV = 1 - [B2IQV3_STRPS] B2IM98 46.54 14 535 123.63 CTP synthase OS =Streptococcus Intracellular pneumoniae (strain CGSP14) GN = pyrG PE = 3SV = 1 - [B2IM98_STRPS] B2IQ39 47.38 9 344 123.25 Uncharacterizedprotein Secretory OS = Streptococcus pneumoniae (released) (strainCGSP14) GN = SPCG_1211 (with PE = 4 SV = 1 - [B2IQ39_STRPS] CS) B2IP0065.99 14 397 123.01 Serine protease OS = Streptococcus N- pneumoniae(strain CGSP14) terminally GN = sphtra PE = 4 SV = 1 - anchored[B2IP00_STRPS] (No CS) B2IN41 35.2 16 429 122.13 Histidine--tRNA ligaseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = hisS PE= 3 SV = 1 - [SYH_STRPS] B2IRW4 40.6 15 436 120.41 GTPase Der OS =Streptococcus Intracellular pneumoniae (strain CGSP14) GN = der PE = 3SV = 1 - [DER_STRPS] B2IR68 35.97 13 531 119.86 Uncharacterized proteinMulti- OS = Streptococcus pneumoniae transmembrane (strain CGSP14) GN =SPCG_1535 PE = 4 SV = 1 - [B2IR68_STRPS] B2IPR2 27.94 18 841 118.97 DNAgyrase subunit A Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = gyrA PE = 3 SV = 1 - [B2IPR2_STRPS] B2IRA1 44.08 17 524118.46 DEAD-box ATP-dependent RNA Intracellular helicase CshA OS =Streptococcus pneumoniae (strain CGSP14) GN = cshA PE = 3 SV = 1 -[B2IRA1_STRPS] B2IM46 55.94 11 345 117.68 Uncharacterized protein N- OS= Streptococcus pneumoniae terminally (strain CGSP14) GN = SPCG_1931anchored PE = 4 SV = 1 - [B2IM46_STRPS] (No CS) B2ILR7 48.44 16 481117.48 6-phosphogluconate Intracellular dehydrogenase, decarboxylatingOS = Streptococcus pneumoniae (strain CGSP14) GN = gnd PE = 3 SV = 1 -[B2ILR7_STRPS] B2IR57 62.02 19 466 116.97 Cof familyprotein/peptidyl-prolyl Intracellular cis-trans isomerase, cyclophilintype OS = Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1524 PE = 4SV = 1 - [B2IR57_STRPS] B2IND9 76.09 12 230 114.4 Cell division ABCtransporter, ATP- Intracellular binding protein FtsE OS = Streptococcuspneumoniae (strain CGSP14) GN = ftsE PE = 4 SV = 1 - [B2IND9_STRPS]B2IPU7 45 13 320 113.45 Ribonucleoside-diphosphate Intracellularreductase subunit beta OS = Streptococcus pneumoniae (strain CGSP14) GN= nrdF PE = 3 SV = 1 - [B2IPU7_STRPS] B2IMS9 43.58 20 553 112.79Ribonuclease J OS = Streptococcus Intracellular pneumoniae (strainCGSP14) GN = rnj PE = 3 SV = 1 - [B2IMS9_STRPS] B2ISQ9 52.02 15 494111.9 UPF0371 protein SPCG_0344 Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_0344 PE = 3 SV = 1 - [Y344_STRPS]B2IPF4 53.38 14 311 110.86 Adhesion lipoprotein Lipid OS = Streptococcuspneumoniae anchored (strain CGSP14) GN = lmb PE = 3 SV = 1 -[B2IPF4_STRPS] B2IM45 50.82 12 427 110.79 UDP-N-acetylglucosamine 1-Intracellular carboxyvinyltransferase OS = Streptococcus pneumoniae(strain CGSP14) GN = murA PE = 3 SV = 1 - [B2IM45_STRPS] B2INN5 58.46 10272 109.88 Cell shape-determining protein N- MreC OS = Streptococcusterminally pneumoniae (strain CGSP14) anchored GN = mreC PE = 3 SV = 1 -(No CS) [B2INN5_STRPS] B2IM15 46.84 10 158 109.73 Uncharacterizedprotein Multi- OS = Streptococcus pneumoniae transmembrane (strainCGSP14) GN = SPCG_1901 PE = 4 SV = 1 - [B2IM15_STRPS] B2IRP9 25.68 14740 109.17 GTP pyrophosphokinase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = relA PE = 3 SV = 1 - [B2IRP9_STRPS]B2INE1 34.71 11 726 108.73 PTS system, IIABC components Multi- OS =Streptococcus pneumoniae transmembrane (strain CGSP14) GN = ptsG PE = 4SV = 1 - [B2INE1_STRPS] B2IST7 48.44 12 417 108.26 Capsularpolysaccharide Intracellular biosynthesis protein, putative OS =Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1812 PE = 3 SV = 1 -[B2IST7_STRPS] B2IP76 71.33 14 286 106.74 Polyamineaminopropyltransferase Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = speE PE = 3 SV = 1 - [B2IP76_STRPS] B2IS23 34.49 22954 105.34 UvrABC system protein A Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = uvrA PE = 3 SV = 1 - [B2IS23_STRPS]B2IS53 52.81 7 89 105.15 30S ribosomal protein S14 Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = rpsN PE = 3 SV = 1 -[RS14_STRPS] B2IQN9 49.79 20 486 105.03 Nicotinate Intracellularphosphoribosyltransferase OS = Streptococcus pneumoniae (strain CGSP14)GN = SPCG_1408 PE = 3 SV = 1 - [B2IQN9_STRPS] B2ISZ6 49.88 13 419 104.34Sugar ABC transporter, sugar- Lipid binding protein OS = Streptococcusanchored pneumoniae (strain CGSP14) GN = SPCG_1871 PE = 4 SV = 1 -[B2ISZ6_STRPS] B2IMA9 47.02 17 487 101.63 Type Irestriction-modification Intracellular system, M subunit OS =Streptococcus pneumoniae (strain CGSP14) GN = hsdM PE = 4 SV = 1 -[B2IMA9_STRPS] B2IQH2 72.19 13 187 100.73 50S ribosomal protein L10Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rplJ PE= 3 SV = 1 - [B2IQH2_STRPS] B2IR61 39.82 14 447 100.57 Asparagine--tRNAligase Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =asnS PE = 3 SV = 1 - [B2IR61_STRPS] B2IM30 50 15 338 100.22Transcriptional regulator, putative Secretory OS = Streptococcuspneumoniae (released) (strain CGSP14) GN = lytR PE = 4 SV = 1 - (with[B2IM30_STRPS] CS) B2ISV9 32.29 10 511 99.16 Choline transporter Multi-OS = Streptococcus pneumoniae transmembrane (strain CGSP14) GN =SPCG_1834 PE = 3 SV = 1 - [B2ISV9_STRPS] B2IS51 78.22 12 101 98.15 50Sribosomal protein L24 Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = rplX PE = 3 SV = 1 - [RL24_STRPS] B2IRX0 76.67 11240 97.84 ABC transporter, ATP-binding Intracellular protein OS =Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1687 PE = 4 SV = 1 -[B2IRX0_STRPS] B2ISX8 46.73 11 336 97.53 Non-canonical purine NTPIntracellular pyrophosphatase OS = Streptococcus pneumoniae (strainCGSP14) GN = SPCG_1853 PE = 3 SV = 1 - [B2ISX8_STRPS] B2IN24 45.35 14505 97.46 4-alpha-glucanotransferase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = malM PE = 3 SV = 1 - [B2IN24_STRPS]B2IM61 54.55 12 275 97.4 Purine operon repressor Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = purR PE = 4 SV = 1 -[B2IM61_STRPS] B2ILX0 55.38 16 455 97.1 Acetyl-CoA carboxylaseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = accC PE= 4 SV = 1 - [B2ILX0_STRPS] B2IQ19 35.91 15 763 96.38 DNA helicase OS =Streptococcus Intracellular pneumoniae (strain CGSP14) GN = pcrA PE = 3SV = 1 - [B2IQ19_STRPS] B2IPZ6 69.23 10 91 95.95 DNA-binding protein HUIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = hlpA PE= 3 SV = 1 - [B2IPZ6_STRPS] B2INE5 31.57 12 396 95.45S-adenosylmethionine synthase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = metK PE = 3 SV = 1 - [METK_STRPS] B2IS8228.53 15 659 94.22 Serine/threonine protein kinase Intracellular/ OS =Streptococcus pneumoniae TMH (strain CGSP14) GN = pkn2 PE = 3 start SV =1 - [B2IS82_STRPS] AFTER 60 B2IS65 58.68 10 121 93.19 30S ribosomalprotein S13 Intracellular OS = Streptococcus pneumoniae (strain CGSP14)GN = rpsM PE = 3 SV = 1 - [RS13_STRPS] B2IMI4 35.02 18 731 92.99Penicillin-binding protein 2A N- OS = Streptococcus pneumoniaeterminally (strain CGSP14) GN = pbp2A PE = 4 anchored SV = 1 -[B2IMI4_STRPS] (No CS) B2IRK4 37.05 17 637 92.87 tRNA uridine 5-Intracellular carboxymethylaminomethyl modification enzyme MnmG OS =Streptococcus pneumoniae (strain CGSP14) GN = mnmG PE = 3 SV = 1 -[B2IRK4_STRPS] B2ISW0 69.01 11 242 92.71 Choline transporterIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1835 PE = 4 SV = 1 - [B2ISW0_STRPS] B2INP6 39.59 10 341 92.59Tryptophan--tRNA ligase Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = trpS PE = 3 SV = 1 - [B2INP6_STRPS] B2INC5 39.5 11281 92.15 Uncharacterized protein Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_0692 PE = 4 SV = 1 - [B2INC5_STRPS]B2IP66 36.64 17 715 91.51 Transcriptional regulator, putativeIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_0885 PE = 4 SV = 1 - [B2IP66_STRPS] B2ISY1 46.58 14 541 91.27Dextran glucosidase DexS, putative Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = dexS PE = 4 SV = 1 - [B2ISY1_STRPS]B2ISK7 38.51 11 444 91.26 Aminopeptidase C Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = pepC PE = 4 SV = 1 -[B2ISK7_STRPS] B2INL6 30.75 8 335 91.2 ABC transporter,substrate-binding Lipid protein, putative OS = Streptococcus anchoredpneumoniae (strain CGSP14) GN = SPCG_2165 PE = 4 SV = 1 - [B2INL6_STRPS]B2IQY9 25.83 11 453 90.37 Chromosomal replication initiatorIntracellular protein DnaA OS = Streptococcus pneumoniae (strain CGSP14)GN = dnaA PE = 3 SV = 1 - [DNAA_STRPS] B2IRS8 22.34 13 685 90.12Penicillin-binding protein 2B N- OS = Streptococcus pneumoniaeterminally (strain CGSP14) GN = pbp2B PE = 4 anchored SV = 1 -[B2IRS8_STRPS] (No CS) B2IS56 53.44 8 131 88.46 50S ribosomal proteinL18 Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =rplR PE = 3 SV = 1 - [B2IS56_STRPS] B2IQ12 40.13 9 319 88.41Ribose-phosphate Intracellular pyrophosphokinase OS = Streptococcuspneumoniae (strain CGSP14) GN = prs PE = 3 SV = 1 - [B2IQ12_STRPS]B2IS67 52.73 9 311 88.17 DNA-directed RNA polymerase Intracellularsubunit alpha OS = Streptococcus pneumoniae (strain CGSP14) GN = rpoA PE= 3 SV = 1 - [B2IS67_STRPS] B2IS59 64.38 9 146 88.05 50S ribosomalprotein L15 Intracellular OS = Streptococcus pneumoniae (strain CGSP14)GN = rplO PE = 3 SV = 1 - [RL15_STRPS] B2IQ61 41.52 9 289 87.7 Cholinekinase OS = Streptococcus Intracellular pneumoniae (strain CGSP14) GN =pck PE = 4 SV = 1 - [B2IQ61_STRPS] B2ISY2 24.4 10 705 87.27 TrehalosePTS system, IIABC Multi- components OS = Streptococcus transmembranepneumoniae (strain CGSP14) GN = treP PE = 4 SV = 1 - [B2ISY2_STRPS]B2IMQ3 25.97 12 801 86.8 Phenylalanine--tRNA ligase beta Intracellularsubunit OS = Streptococcus pneumoniae (strain CGSP14) GN = pheT PE = 3SV = 1 - [B2IMQ3_STRPS] B2IQ02 60.82 9 97 86.49 50S ribosomal proteinL27 Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =rpmA PE = 3 SV = 1 - [RL27_STRPS] B2IN58 40.3 11 335 86.4Uncharacterized protein Multi- OS = Streptococcus pneumoniaetransmembrane (strain CGSP14) GN = SPCG_2101 PE = 4 SV = 1 -[B2IN58_STRPS] B2IQW4 62.2 10 209 84.4 Amino acid ABC transporter, ATP-Intracellular binding protein OS = Streptococcus pneumoniae (strainCGSP14) GN = SPCG_1486 PE = 4 SV = 1 - [B2IQW4_STRPS] B2IP74 35.23 13491 84.24 Lysine decarboxylase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = cad PE = 4 SV = 1 - [B2IP74_STRPS]B2IS70 42.49 11 433 83.11 UPF0210 protein SPCG_0246 Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = SPCG_0246 PE = 3 SV = 1 -[B2IS70_STRPS] B2IQ66 64.29 13 280 83.05 LicD2 protein OS =Streptococcus Intracellular pneumoniae (strain CGSP14) GN = licD2 PE = 4SV = 1 - [B2IQ66_STRPS] B2IRK5 34.17 12 559 81.76 Ribonuclease J OS =Streptococcus Intracellular pneumoniae (strain CGSP14) GN = rnj PE = 3SV = 1 - [B2IRK5_STRPS] B2IR84 68.24 12 296 81.42 Nucleotide-bindingprotein Intracellular SPCG_1551 OS = Streptococcus pneumoniae (strainCGSP14) GN = SPCG_1551 PE = 3 SV = 1 - [Y1551_STRPS] B2IMB5 34.93 15 60781.24 Chaperone protein DnaK Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = dnaK PE = 3 SV = 1 - [DNAK_STRPS] B2IQ62 58.09 12346 80.31 Alcohol dehydrogenase, zinc- Intracellular containing OS =Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1234 PE = 4 SV = 1 -[B2IQ62_STRPS] B2IMB9 37.86 11 243 79.94 ABC transporter, ATP-bindingIntracellular protein OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_0494 PE = 4 SV = 1 - [B2IMB9_STRPS] B2IRD4 56.18 5 89 79.7 30Sribosomal protein S15 Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = rpsO PE = 3 SV = 1 - [RS15_STRPS] B2IN68 50.79 8126 79.14 Uncharacterized protein N- OS = Streptococcus pneumoniaeterminally (strain CGSP14) GN = SPCG_0635 anchored PE = 4 SV = 1 -[B2IN68_STRPS] (No CS) B2IQN0 44.09 3 127 78.99 Uncharacterized proteinN- OS = Streptococcus pneumoniae terminally (strain CGSP14) GN =SPCG_1399 anchored PE = 4 SV = 1 - [B2IQN0_STRPS] (No CS) B2ILY0 36.5814 514 78.96 Peptide chain release factor 3 Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = prfC PE = 3 SV = 1 -[RF3_STRPS] B2IS54 57.58 8 132 78.42 30S ribosomal protein S8Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rpsH PE= 3 SV = 1 - [RS8_STRPS] B2IMB6 35.19 13 378 78.27 Chaperone proteinDnaJ Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =dnaJ PE = 3 SV = 1 - [B2IMB6_STRPS] B2IQS5 42.11 12 247 78.16 Amino acidABC transporter, ATP- Intracellular binding protein OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_1447 PE = 4 SV = 1 - [B2IQS5_STRPS]B2IRH0 33.01 12 512 76.84 ABC transporter, substrate-binding Lipidprotein OS = Streptococcus anchored pneumoniae (strain CGSP14) GN =SPCG_0089 PE = 4 SV = 1 - [B2IRH0_STRPS] B2IPW9 50.88 11 283 76.72Ribosome biogenesis GTPase A Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = SPCG_1141 PE = 3 SV = 1 - [B2IPW9_STRPS] B2IN1848.09 11 418 76.44 Tyrosine--tRNA ligase Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = tyrS PE = 3 SV = 1 -[SYY_STRPS] B2IQV9 33.39 13 572 75.14 Phosphoglucomutase IntracellularOS = Streptococcus pneumoniae (strain CGSP14) GN = pgm PE = 3 SV = 1 -[B2IQV9_STRPS] B2IP77 38.9 9 419 74.94 Uncharacterized proteinIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_0896 PE = 4 SV = 1 - [B2IP77_STRPS] B2IQ63 53.19 8 235 74.222-C-methyl-D-erythritol 4- Intracellular phosphate cytidylyltransferaseOS = Streptococcus pneumoniae (strain CGSP14) GN = ispD PE = 3 SV = 1 -[ISPD_STRPS] B2IRQ4 65.34 11 251 74.2 Manganese ABC transporter, ATP-Intracellular binding protein OS = Streptococcus pneumoniae (strainCGSP14) GN = SPCG_1621 PE = 4 SV = 1 - [B2IRQ4_STRPS] B2IQX3 62.36 8 17873.32 ATP synthase subunit delta Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = atpH PE = 3 SV = 1 - [ATPD_STRPS] B2INX427.48 12 826 73.21 DNA topoisomerase 4 subunit A Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = parC PE = 3 SV = 1 -[B2INX4_STRPS] B2IN76 40.67 12 450 72.88 UDP-N-acetylmuramoylalanine--D-Intracellular glutamate ligase OS = Streptococcus pneumoniae (strainCGSP14) GN = murD PE = 3 SV = 1 - [MURD_STRPS] B2IPC3 41.47 10 299 72.75GTPase Era OS = Streptococcus Intracellular pneumoniae (strain CGSP14)GN = era PE = 3 SV = 1 - [ERA_STRPS] B2IRX7 40.79 13 353 72.1 Methionineimport ATP-binding Intracellular protein MetN OS = Streptococcuspneumoniae (strain CGSP14) GN = metN PE = 3 SV = 1 - [B2IRX7_STRPS]B2INU1 50.65 9 306 70.6 Bifunctional protein FolD Lipid OS =Streptococcus pneumoniae anchored (strain CGSP14) GN = folD PE = 3 SV =1 - [B2INU1_STRPS] B2ILW4 36.11 6 324 69.62 Enoyl-(Acyl-carrier-protein)Intracellular reductase OS = Streptococcus pneumoniae (strain CGSP14) GN= fabK PE = 4 SV = 1 - [B2ILW4_STRPS] B2IS45 40.35 3 114 69.56 50Sribosomal protein L22 Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = rplV PE = 3 SV = 1 - [RL22_STRPS] B2IPZ1 30.21 13652 67.59 DNA ligase OS = Streptococcus Intracellular pneumoniae (strainCGSP14) GN = ligA PE = 3 SV = 1 - [DNLJ_STRPS] B2IMZ3 34.66 12 603 66.96ABC transporter, ATP- Multi- binding/permease protein transmembrane OS =Streptococcus pneumoniae (strain CGSP14) GN = SPCG_2040 PE = 4 SV = 1 -[B2IMZ3_STRPS] B2IS63 54.72 6 212 66.76 Adenylate kinase OS =Streptococcus Intracellular pneumoniae (strain CGSP14) GN = adk PE = 3SV = 1 - [KAD_STRPS] B2INV5 53.01 6 83 65.46 30S ribosomal protein S20Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rpsT PE= 3 SV = 1 - [B2INV5_STRPS] B2IRK2 38.07 10 373 65.45 tRNA-specific2-thiouridylase Intracellular MnmA OS = Streptococcus pneumoniae (strainCGSP14) GN = mnmA PE = 3 SV = 2 - [MNMA_STRPS] B2IRB7 27.22 2 158 65.21Uncharacterized protein N- OS = Streptococcus pneumoniae terminally(strain CGSP14) GN = SPCG_1584 anchored PE = 4 SV = 1 - [B2IRB7_STRPS](No CS) B2INE6 56.27 10 311 64.68 Dihydroorotate dehydrogenaseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = pyrD PE= 3 SV = 1 - [B2INE6_STRPS] B2IMB0 27.41 15 777 64.13 Type Irestriction-modification Intracellular system, R subunit OS =Streptococcus pneumoniae (strain CGSP14) GN = hsdR PE = 4 SV = 1 -[B2IMB0_STRPS] B2ISQ4 20.27 11 750 63.95 Penicillin-binding protein 2XN- OS = Streptococcus pneumoniae terminally (strain CGSP14) GN = pbp2XPE = 4 anchored SV = 1 - [B2ISQ4_STRPS] (No CS) B2IPQ3 27.88 12 55663.73 Formate--tetrahydrofolate ligase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = fhs PE = 3 SV = 1 - [FTHS_STRPS] B2IMU741.84 5 141 63.38 50S ribosomal protein L11 Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = rplK PE = 3 SV = 1 -[RL11_STRPS] B2ISY0 38 4 100 62.82 UPF0154 protein SPCG_1855 N- OS =Streptococcus pneumoniae terminally (strain CGSP14) GN = SPCG_1855anchored PE = 3 SV = 1 - [B2ISY0_STRPS] (No CS) B2IQP4 29.49 9 434 62.33Peptidase, U32 family Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = SPCG_1416 PE = 4 SV = 1 - [B2IQP4_STRPS] B2IR7957.14 11 259 62.31 Uncharacterized protein N- OS = Streptococcuspneumoniae terminally (strain CGSP14) GN = SPCG_1546 anchored PE = 4 SV= 1 - [B2IR79_STRPS] (No CS) B2ILW2 21.6 8 324 62.193-oxoacyl-[acyl-carrier-protein] Intracellular synthase 3 OS =Streptococcus pneumoniae (strain CGSP14) GN = fabH PE = 3 SV = 1 -[FABH_STRPS] B2IPZ9 43.28 9 305 61.07 Riboflavin biosynthesis proteinIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = mreA PE= 3 SV = 1 - [B2IPZ9_STRPS] B2IN32 29.98 13 587 60.72 Aspartate--tRNAligase Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =aspS PE = 3 SV = 1 - [SYD_STRPS] B2IRR9 42.46 7 179 60.72 Cell divisionprotein SepF Intracellular OS = Streptococcus pneumoniae (strain CGSP14)GN = sepF PE = 3 SV = 1 - [SEPF_STRPS] B2IN19 28.12 13 690 60.07Cation-transporting ATPase, EI-E2 Intracellular family OS =Streptococcus pneumoniae (strain CGSP14) GN = ctpC PE = 3 SV = 1 -[B2IN19_STRPS] B2IQY8 36.71 9 444 59.95 UDP-N-acetylmuramate--L-alanineIntracellular ligase OS = Streptococcus pneumoniae (strain CGSP14) GN =murC PE = 3 SV = 1 - [MURC_STRPS] B2IS26 33.86 8 189 59.81Uncharacterized protein Lipid OS = Streptococcus pneumoniae anchored(strain CGSP14) GN = SPCG_0204 PE = 4 SV = 1 - [B2IS26_STRPS] B2INY745.25 8 263 59.27 ABC transporter, ATP-binding Intracellular protein OS= Streptococcus pneumoniae (strain CGSP14) GN = SPCG_0812 PE = 4 SV =1 - [B2INY7_STRPS] B2ILR6 14.66 6 464 58.78 Mid-cell-anchored protein ZN- OS = Streptococcus pneumoniae terminally (strain CGSP14) GN = mapZ PE= 3 anchored SV = 1 - [B2ILR6_STRPS] (No CS) B2ILX8 36.68 10 488 58.59Glutamyl-tRNA(Gln) Intracellular amidotransferase subunit A OS =Streptococcus pneumoniae (strain CGSP14) GN = gatA PE = 3 SV = 1 -[GATA_STRPS] B2IMN3 35.26 7 380 58.5 Queuine tRNA-ribosyltransferaseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = tgt PE= 3 SV = 1 - [TGT_STRPS] B2IN17 21.09 12 825 58.4 Penicillin-bindingprotein 1B Intracellular/ OS = Streptococcus pneumoniae TMH (strainCGSP14) GN = pbp1B PE = 4 start SV = 1 - [B2IN17_STRPS] AFTER 60 B2IR2745.45 8 330 57.61 Phosphate acyltransferase Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = plsX PE = 3 SV = 1 -[PLSX_STRPS] B2INF7 73.21 6 112 56.77 ATP cone domain-containingIntracellular protein OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_0728 PE = 4 SV = 1 - [B2INF7_STRPS] B2IMZ8 20.78 9 563 56.13Arginine--tRNA ligase Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = argS PE = 3 SV = 1 - [SYR_STRPS] B2ILU5 35.6 10 42755.96 Trigger factor OS = Streptococcus Intracellular pneumoniae (strainCGSP14) GN = tig PE = 3 SV = 1 - [TIG_STRPS] B2INK8 56.55 9 336 55.86tRNA-dihydrouridine synthase Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = SPCG_2157 PE = 3 SV = 1 - [B2INK8_STRPS] B2IM9037.14 11 560 55.51 ABC transporter, ATP-binding Intracellular protein OS= Streptococcus pneumoniae (strain CGSP14) GN = SPCG_0465 PE = 4 SV =1 - [B2IM90_STRPS] B2IQA4 37.72 12 448 54.94 Glutamate dehydrogenaseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = gdhA PE= 3 SV = 1 - [B2IQA4_STRPS] B2IPP0 30.1 8 495 54.8 Glucose-6-phosphate1- Intracellular dehydrogenase OS = Streptococcus pneumoniae (strainCGSP14) GN = zwf PE = 3 SV = 1 - [B2IPP0_STRPS] B2IPZ5 24.33 13 63354.29 ABC transporter, ATP-binding Intracellular protein OS =Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1167 PE = 4 SV = 1 -[B2IPZ5_STRPS] B2IS10 23.73 8 649 54.27 DNA mismatch repair protein MutLIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = mutL PE= 3 SV = 1 - [MUTL_STRPS] B2IN96 47.62 10 378 54.14 Lactate oxidase OS =Streptococcus Intracellular pneumoniae (strain CGSP14) GN = lctO PE = 4SV = 1 - [B2IN96_STRPS] B2IND6 51.69 6 236 53.89 Branched-chain aminoacid ABC Intracellular transporter, ATP-binding protein OS =Streptococcus pneumoniae (strain CGSP14) GN = SPCG_0703 PE = 4 SV = 1 -[B2IND6_STRPS] B2IS98 58.85 9 209 53.7 Probable nicotinate-nucleotideIntracellular adenylyltransferase OS = Streptococcus pneumoniae (strainCGSP14) GN = nadD PE = 3 SV = 1 - [B2IS98_STRPS] B2IPN8 43.01 8 27253.21 Cof family protein Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = SPCG_1058 PE = 4 SV = 1 - [B2IPN8_STRPS] B2INN851.27 10 275 52.83 Energy-coupling factor transporter N- ATP-bindingprotein EcfA terminally OS = Streptococcus pneumoniae anchored (strainCGSP14) GN = ecfA PE = 3 SV = 1 - (No CS) [B2INN8_STRPS] B2ISR0 23.55 8535 52.43 Glucan 1,6-alpha-glucosidase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = dexB PE = 4 SV = 1 - [B2ISR0_STRPS]B2IRF6 52.19 8 228 52.22 Potassium uptake protein, Trk N- family OS =Streptococcus terminally pneumoniae (strain CGSP14) anchored GN = trkAPE = 4 SV = 1 - (with CS) [B2IRF6_STRPS] B2IM43 17.16 7 443 51.63 CBSdomain protein Multi- OS = Streptococcus pneumoniae transmembrane(strain CGSP14) GN = SPCG_1928 PE = 4 SV = 1 - [B2IM43_STRPS] B2ILX2 406 255 51.58 Acetyl-coenzyme A carboxylase Intracellular carboxyltransferase subunit alpha OS = Streptococcus pneumoniae (strain CGSP14)GN = accA PE = 3 SV = 1 - [ACCA_STRPS] B2ISA0 34.78 10 368 51.32GTP-binding protein Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = SPCG_1723 PE = 4 SV = 1 - [B2ISA0_STRPS] B2IPB9 23.19 10677 50.7 Endo-beta-N-acetylglucosaminidase Secretory OS = Streptococcuspneumoniae (released) (strain CGSP14) GN = lytB PE = 4 SV = 1 - (with[B2IPB9_STRPS] CS) B2IQW9 61.87 6 139 50.69 ATP synthase epsilon chainIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = atpC PE= 3 SV = 1 - [ATPE_STRPS] B2IPN9 24.94 7 429 50.34 Signal recognitionparticle receptor Intracellular FtsY OS = Streptococcus pneumoniae(strain CGSP14) GN = ftsY PE = 3 SV = 1 - [B2IPN9_STRPS] B2IRI7 17.95 9713 49.57 Uncharacterized protein Multi- OS = Streptococcus pneumoniaetransmembrane (strain CGSP14) GN = SPCG_0106 PE = 4 SV = 1 -[B2IRI7_STRPS] B2IQH3 24.39 7 488 49.39 Chlorohydrolase OS =Streptococcus Intracellular pneumoniae (strain CGSP14) GN = trzA PE = 4SV = 1 - [B2IQH3_STRPS] B2IRD2 21.59 7 264 48.68 Acyltransferase familyprotein Multi- OS = Streptococcus pneumoniae transmembrane (strainCGSP14) GN = SPCG_1599 PE = 4 SV = 1 - [B2IRD2_STRPS] B2IR63 31.14 7 39548.62 Aspartate aminotransferase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = aspB PE = 3 SV = 1 - [B2IR63_STRPS]B2INK4 23.36 10 608 48.46 Uncharacterized protein N- OS = Streptococcuspneumoniae terminally (strain CGSP14) GN = SPCG_2153 anchored PE = 4 SV= 1 - [B2INK4_STRPS] (No CS) B2INI8 28.14 7 501 48.06 Zinc ABCtransporter, zinc-binding Lipid lipoprotein OS = Streptococcus anchoredpneumoniae (strain CGSP14) GN = SPCG_2137 PE = 3 SV = 1 - [B2INI8_STRPS]B2IPU4 40.71 7 253 47.82 Lactose phosphotransferase system Intracellularrepressor OS = Streptococcus pneumoniae (strain CGSP14) GN = lacR PE = 4SV = 1 - [B2IPU4_STRPS] B2INN7 41.94 8 279 47.56 Energy-coupling factortransporter N- ATP-binding protein EcfA terminally OS = Streptococcuspneumoniae anchored (strain CGSP14) GN = ecfA PE = 3 SV = 1 - (No CS)[B2INN7_STRPS] B2IS50 52.46 7 122 47.46 50S ribosomal protein L14Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rplN PE= 3 SV = 1 - [RL14_STRPS] B2IRI6 32.63 3 95 47.46 Bacteriocin, putativeN- OS = Streptococcus pneumoniae terminally (strain CGSP14) GN =SPCG_0105 anchored PE = 4 SV = 1 - [B2IRI6_STRPS] (with CS) B2IPJ1 50 9264 47.22 Iron-compound ABC transporter, Intracellular ATP-bindingprotein OS = Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1013 PE= 4 SV = 1 - [B2IPJ1_STRPS] B2IPR3 40.89 7 247 47.21 Uncharacterizedprotein N- OS = Streptococcus pneumoniae terminally (strain CGSP14) GN =SPCG_1083 anchored PE = 4 SV = 1 - [B2IPR3_STRPS] (with CS) B2IR24 28.289 389 47.15 Uncharacterized protein Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_0036 PE = 3 SV = 1 - [B2IR24_STRPS]B2ISX7 60.12 8 173 47.03 Phosphoesterase OS = StreptococcusIntracellular pneumoniae (strain CGSP14) GN = SPCG_1852 PE = 3 SV = 1 -[B2ISX7_STRPS] B2IM86 29.62 8 449 46.75 Potassium uptake protein, Trk N-family OS = Streptococcus terminally pneumoniae (strain CGSP14) anchoredGN = trkA PE = 4 SV = 1 - (with CS) [B2IM86_STRPS] B2IPT8 33.13 9 32646.7 Tagatose 1,6-diphosphate aldolase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = lacD PE = 3 SV = 1 - [B2IPT8_STRPS]B2ILZ5 39.81 10 540 46.59 60 kDa chaperonin Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = groL PE = 3 SV = 1 -[CH60_STRPS] B2ILX7 34.58 8 480 46.43 Aspartyl/glutamyl-tRNA(Asn/Gln)Intracellular amidotransferase subunit B OS = Streptococcus pneumoniae(strain CGSP14) GN = gatB PE = 3 SV = 1 - [GATB_STRPS] B2IMQ9 16.96 8737 45.59 Polyribonucleotide Intracellular nucleotidyltransferase OS =Streptococcus pneumoniae (strain CGSP14) GN = pnp PE = 3 SV = 2 -[PNP_STRPS] B2IQN3 45.45 5 66 44.91 30S ribosomal protein S21Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rpsU PE= 3 SV = 1 - [B2IQN3_STRPS] B2IR90 38.37 2 172 44.68 Non-hemeiron-containing ferritin Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = SPCG_1557 PE = 3 SV = 1 - [B2IR90_STRPS] B2IMS242.66 7 293 44.59 Fructose-bisphosphate aldolase Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = fba PE = 3 SV = 1 -[B2IMS2_STRPS] B2IRX2 10.53 5 399 44.06 Uncharacterized protein Multi-OS = Streptococcus pneumoniae transmembrane (strain CGSP14) GN =SPCG_1689 PE = 4 SV = 1 - [B2IRX2_STRPS] B2ISH7 20.98 10 815 43.34Formate acetyltransferase, putative Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = pflF PE = 4 SV = 1 - [B2ISH7_STRPS]B2ISG1 26.84 7 190 43.06 Uncharacterized protein Multi- OS =Streptococcus pneumoniae transmembrane (strain CGSP14) GN = SPCG_1779 PE= 4 SV = 1 - [B2ISG1_STRPS] B2IMY2 73.21 7 56 42.86 Uncharacterizedprotein Secretory OS = Streptococcus pneumoniae (released) (strainCGSP14) GN = SPCG_0626 (with PE = 4 SV = 1 - [B2IMY2_STRPS] CS) B2IM3615.5 8 671 42.81 Uncharacterized protein Multi- OS = Streptococcuspneumoniae transmembrane (strain CGSP14) GN = SPCG_1921 PE = 4 SV = 1 -[B2IM36_STRPS] B2IM12 26.96 8 471 42.77 Pneumolysin OS = StreptococcusIntracellular pneumoniae (strain CGSP14) GN = ply PE = 4 SV = 1 -[B2IM12_STRPS] B2INM4 34.62 6 182 42.71 Ribosomal subunit interfaceprotein Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_2173 PE = 4 SV = 1 - [B2INM4_STRPS] B2IPW4 27.34 9 567 42.61Dihydrolipoyl dehydrogenase Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = acoL PE = 4 SV = 1 - [B2IPW4_STRPS] B2IQ29 38.89 6180 42.56 Hypoxanthine-guanine Intracellular phosphoribosyltransferaseOS = Streptococcus pneumoniae (strain CGSP14) GN = hgt PE = 4 SV = 1 -[B2IQZ9_STRPS] B2IS21 19.23 6 234 42.44 Uncharacterized protein Multi-OS = Streptococcus pneumoniae transmembrane (strain CGSP14) GN =SPCG_0199 PE = 4 SV = 1 - [B2IS21_STRPS] B2IPB4 18.18 3 66 42.39 50Sribosomal protein L35 Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = rpmI PE = 3 SV = 1 - [RL35_STRPS] B2IM29 23.68 5418 42.24 Putative competence-damage Intracellular inducible protein OS= Streptococcus pneumoniae (strain CGSP14) GN = cinA PE = 3 SV = 1 -[CINA_STRPS] B2IR86 66.5 11 197 41.89 Probable GTP-binding protein EngBIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = engB PE= 3 SV = 1 - [ENGB_STRPS] B2IPS2 46.7 4 212 41.29 Uridine kinase OS =Streptococcus Intracellular pneumoniae (strain CGSP14) GN = udk PE = 3SV = 1 - [URK_STRPS] B2INP0 22.46 5 276 41.12 Uncharacterized proteinIntracellular/ OS = Streptococcus pneumoniae TMH (strain CGSP14) GN =SPCG_2189 start PE = 4 SV = 1 - [B2INP0_STRPS] AFTER 60 B2ILX1 42.71 7288 41.03 Acetyl-coenzyme A carboxylase Intracellular carboxyltransferase subunit beta OS = Streptococcus pneumoniae (strain CGSP14)GN = accD PE = 3 SV = 1 - [B2ILX1_STRPS] B2IMY9 31.48 10 486 40.7Glutamate--tRNA ligase Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = gltX PE = 3 SV = 1 - [SYE_STRPS] B2ISL4 51.11 9 27040.64 Cof family protein Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = SPCG_0299 PE = 4 SV = 1 - [B2ISL4_STRPS] B2IQH742.21 5 289 40.55 Homoserine kinase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = thrB PE = 3 SV = 1 - [KHSE_STRPS] B2IS8936.54 7 208 40.54 Guanylate kinase OS = Streptococcus Intracellularpneumoniae (strain CGSP14) GN = gmk PE = 3 SV = 1 - [B2IS89_STRPS]B2IRQ3 18.92 7 650 40.16 Endopeptidase O OS = StreptococcusIntracellular pneumoniae (strain CGSP14) GN = pepO PE = 4 SV = 1 -[B2IRQ3_STRPS] B2IR80 26.03 6 292 39.76 Uncharacterized protein Multi-OS = Streptococcus pneumoniae transmembrane (strain CGSP14) GN =SPCG_1547 PE = 4 SV = 1 - [B2IR80_STRPS] B2INZ2 20.82 6 413 39.4D-alanyl-D-alanine N- carboxypeptidase terminally OS = Streptococcuspneumoniae anchored (strain CGSP14) GN = dacA PE = 3 (with CS) SV = 1 -[B2INZ2_STRPS] B2IR93 45.95 6 259 39.26 Triosephosphate isomeraseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = tpiA PE= 3 SV = 1 - [B2IR93_STRPS] B2IP56 34.48 8 290 39.23 Uncharacterizedprotein Lipid OS = Streptococcus pneumoniae anchored (strain CGSP14) GN= SPCG_0875 PE = 4 SV = 1 - [B2IP56_STRPS] B2IPZ7 46.59 6 279 39.17 DegVfamily protein Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = SPCG_1169 PE = 4 SV = 1 - [B2IPZ7_STRPS] B2INK5 19.52 8 50239.03 Glycerol kinase OS = Streptococcus Intracellular pneumoniae(strain CGSP14) GN = glpK PE = 3 SV = 1 - [GLPK_STRPS] B2INM2 32.67 7150 38.84 50S ribosomal protein L9 Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = rplI PE = 3 SV = 1 - [RL9_STRPS] B2IPH621.17 4 359 38.78 Peptide chain release factor 1 Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = prfA PE = 3 SV = 1 -[RF1_STRPS] B2ING0 44.21 3 95 38.52 30S ribosomal protein S16Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rpsP PE= 3 SV = 1 - [RS16_STRPS] B2IQZ0 33.07 7 378 38.34 DNA polymerase IIIsubunit beta Intracellular OS = Streptococcus pneumoniae (strain CGSP14)GN = dnaN PE = 4 SV = 1 - [B2IQZ0_STRPS] B2IR11 29.29 8 420 37.79 DNArepair protein radA Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = radA PE = 3 SV = 1 - [B2IR11_STRPS] B2ISK0 7.86 8 1463 37.3DNA polymerase III PolC-type Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = polC PE = 3 SV = 1 - [B2ISK0_STRPS] B2IMD8 40.15 8264 37.04 Uncharacterized protein Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_0513 PE = 4 SV = 1 - [B2IMD8_STRPS]B2IP06 33.66 7 303 36.91 Tagatose-6-phosphate kinase Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = fruB PE = 3 SV = 1 -[B2IP06_STRPS] B2IPT6 46.2 5 171 36.86 Galactose-6-phosphate isomeraseIntracellular subunit LacB OS = Streptococcus pneumoniae (strain CGSP14)GN = lacB PE = 3 SV = 1 - [LACB_STRPS] B2IMT1 28.33 7 406 36.74Beta-lactam resistance factor Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = murM PE = 4 SV = 1 - [B2IMT1_STRPS]B2IN52 24.87 8 567 36.72 Dihydroxy-acid dehydratase Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = ilvD PE = 3 SV = 1 -[ILVD_STRPS] B2IRH4 23.13 5 320 36.59 Uncharacterized protein N- OS =Streptococcus pneumoniae terminally (strain CGSP14) GN = SPCG_0093anchored PE = 4 SV = 1 - [B2IRH4_STRPS] (No CS) B2IME1 27.51 7 378 36.3Transcription Intracellular termination/antitermination protein NusA OS= Streptococcus pneumoniae (strain CGSP14) GN = nusA PE = 3 SV = 1 -[B2IME1_STRPS] B2ISX2 51.67 9 240 36.23 Pseudouridine synthaseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rluB PE= 3 SV = 1 - [B2ISX2_STRP5] B2IN61 77.55 6 49 35.89 50S ribosomalprotein L33 Intracellular OS = Streptococcus pneumoniae (strain CGSP14)GN = rpmG PE = 3 SV = 1 - [RL33_STRPS] B2IQ98 60.64 4 94 35.66 50Sribosomal protein L31 type B Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = rpmE PE = 3 SV = 1 - [B2IQ98_STRPS] B2IPY5 32.19 7379 35.53 Glycogen biosynthesis protein GlgD Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = glgD PE = 4 SV = 1 -[B2IPY5_STRPS] B2IRA3 21.23 4 438 35.35 Pyridine nucleotide-disulfideIntracellular oxidoreductase OS = Streptococcus pneumoniae (strainCGSP14) GN = merA PE = 3 SV = 1 - [B2IRA3 STRPS] B2IR65 48.8 6 209 35.25Uncharacterized protein Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = SPCG_1532 PE = 4 SV = 1 - [B2IR65_STRPS] B2INN136.99 7 346 34.94 Elongation factor Ts Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = tsf PE = 3 SV = 1 - [EFTS_STRPS] B2IRV519.83 3 343 34.85 Phospho-2-dehydro-3- Intracellular deoxyheptonatealdolase OS = Streptococcus pneumoniae (strain CGSP14) GN = aroF PE = 3SV = 1 - [B2IRV5_STRPS] B2IN60 53.33 4 60 34.75 50S ribosomal proteinL32 Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =rpmF PE = 3 SV = 1 - [RL32_STRPS] B2IQ13 32.08 6 371 34.73Aminotransferase, class-V Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = SPCG_1185 PE = 3 SV = 1 - [B2IQ13_STRPS] B2INQ136.11 5 180 34.11 Transcriptional regulator, TetR Intracellular familyOS = Streptococcus pneumoniae (strain CGSP14) GN = SPCG_2200 PE = 4 SV =1 - [B2INQ1_STRPS] B2IP08 15.91 7 767 33.57 SpoE family protein Multi-OS = Streptococcus pneumoniae transmembrane (strain CGSP14) GN = ftsK PE= 3 SV = 1 - [B2IP08_STRPS] B2IRW8 41.24 6 177 33.56 Transcriptionalrepressor NrdR Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = nrdR PE = 3 SV = 1 - [B2IRW8_STRPS] B2ISL9 57.61 6 18433.49 GTP cyclohydrolase 1 Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = folE PE = 3 SV = 1 - [GCH1_STRPS] B2IQL2 25.35 4217 32.6 Phosphate-specific transport Intracellular system accessoryprotein PhoU OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1384 PE = 3 SV = 1 - [B2IQL2_STRPS] B2ILW1 39.58 4 144 32.47Transcriptional regulator, MarR Intracellular family OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_0414 PE = 4 SV = 1 - [B2ILW1_STRPS]B2IS33 15.25 5 518 32.44 Competence-induced protein Ccs4 Multi- OS =Streptococcus pneumoniae transmembrane (strain CGSP14) GN = SPCG_0211 PE= 4 SV = 1 - [B2IS33_STRPS] B2INX7 16.53 4 236 32.43 Uncharacterizedprotein Multi- OS = Streptococcus pneumoniae transmembrane (strainCGSP14) GN = SPCG_0802 PE = 4 SV = 1 - [B2INX7_STRPS] B2INM3 14.16 7 65732.29 DHH subfamily 1 protein Multi- OS = Streptococcus pneumoniaetransmembrane (strain CGSP14) GN = SPCG_2172 PE = 4 SV = 1 -[B2INM3_STRPS] B2INY8 22.62 6 420 32.18 Uncharacterized proteinIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_0813 PE = 4 SV = 1 - [B2INY8_STRPS] B2IN02 20.77 5 443 32.03 Sensorhistidine kinase PnpS Multi- OS = Streptococcus pneumoniae transmembrane(strain CGSP14) GN = SPCG_2049 PE = 4 SV = 1 - [B2IN02_STRPS] B2IMR813.73 5 459 31.87 Transmembrane protein Vexp3 Multi- OS = Streptococcuspneumoniae transmembrane (strain CGSP14) GN = SPCG_0562 PE = 3 SV = 1 -[B2IMR8_STRPS] B2IP11 34.65 7 404 31.83 Probable tRNA sulfurtransferaseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = thil PE= 3 SV = 1 - [THII_STRPS] B2INZ1 26.38 8 470 31.82 FeS assembly proteinSufB Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_0816 PE = 4 SV = 1 - [B2INZ1_STRPS] B2IQ37 41.27 5 252 31.78 ABCtransporter, ATP-binding Intracellular protein OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_1209 PE = 4 SV = 1 - [B2IQ37_STRPS]B2IN78 31.83 6 399 31.49 Cell division protein DivlB Intracellular/ OS =Streptococcus pneumoniae TMH (strain CGSP14) GN = divlB PE = 3 start SV= 1 - [B2IN78_STRPS] AFTER 60 B2IS71 21.69 5 249 31.41 Phosphoglyceratemutase family Intracellular protein OS = Streptococcus pneumoniae(strain CGSP14) GN = gpmB PE = 4 SV = 1 - [B2IS71_STRPS] B2IQJ2 23.71 5388 31.39 Chorismate synthase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = aroC PE = 3 SV = 1 - [AROC_STRPS] B2IST913.89 7 583 31.29 ABC transporter, ATP- Multi- binding/permease proteintransmembrane OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1814 PE = 4 SV = 1 - [B2IST9_STRPS] B2IPQ2 32.05 6 234 30.92Uncharacterized protein Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = SPCG_1072 PE = 4 SV = 1 - [B2IPQ2_STRPS] B2ISM018.89 3 270 30.69 7,8-dihydroneopterin aldolase Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = sulD PE = 3 SV = 1 -[B2ISM0_STRPS] B2IMH1 16.74 6 472 30.67 Cof family protein IntracellularOS = Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1962 PE = 4 SV =1 - [B2IMH1_STRPS] B2ISV2 17.09 5 392 30.34 Galactokinase OS =Streptococcus Intracellular pneumoniae (strain CGSP14) GN = galK PE = 3SV = 1 - [B2ISV2_STRPS] B2IR20 12.6 8 889 30.19 DNA polymerase OS =Streptococcus Intracellular pneumoniae (strain CGSP14) GN = polA PE = 3SV = 1 - [B2IR20_STRPS] B2IR50 26.4 6 481 30UDP-N-acetylmuramoyl-L-alanyl-D- Intracellular glutamate--L-lysineligase OS = Streptococcus pneumoniae (strain CGSP14) GN = murE PE = 3 SV= 1 - [B2IR50_STRPS] B2ISI6 20.93 6 258 29.94 Isoprenyl transferaseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = uppS PE= 3 SV = 1 - [B2ISI6_STRPS] B2IMQ1 31.47 7 375 29.85 Phenylalanine--tRNAligase alpha Intracellular subunit OS = Streptococcus pneumoniae (strainCGSP14) GN = pheS PE = 3 SV = 1 - [B2IMQI_STRPS] B2ISS1 20.39 6 40729.77 Tryptophan synthase beta chain Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = trpB PE = 3 SV = 1 - [TRPB_STRPS] B2IMZ022.05 5 449 29.65 Glucose-6-phosphate isomerase Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = pgi PE = 3 SV = 1 -[G6PI_STRPS] B2IRD1 13.75 4 778 29.59 Cation-transporting ATPase, E1-E2Multi- family OS = Streptococcus transmembrane pneumoniae (strainCGSP14) GN = ctpE PE = 3 SV = 1 - [B2IRD1_STRPS] B2IQ17 51.09 5 22929.49 Glutamine amidotransferase, class I Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1189 PE = 4 SV = 1 -[B2IQ17_STRPS] B2IPV5 30.5 5 200 29.44 Uncharacterized proteinIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1127 PE = 4 SV = 1 - [B2IPV5_STRPS] B2ILW5 27.78 6 306 29.34Malonyl CoA-acyl carrier protein Intracellular transacylase OS =Streptococcus pneumoniae (strain CGSP14) GN = fabD PE = 3 SV = 1 -[B2ILW5_STRPS] B2IQI6 29.48 6 424 29.26 Psr protein OS = StreptococcusIntracellular/ pneumoniae (strain CGSP14) TMH GN = SPCG_1358 PE = 4 SV =1 - start [B2IQI6_STRPS] AFTER 60 B2IPB3 40.51 5 195 28.99 Translationinitiation factor IF-3 Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = infC PE = 3 SV = 1 - [B2IPB3_STRPS] B2IM63 18.52 7432 28.92 Competence-induced protein Ccs50 N- OS = Streptococcuspneumoniae terminally (strain CGSP14) GN = SPCG_1944 anchored PE = 4 SV= 1 - [B2IM63_STRPS] (No CS) B2IP99 44.53 6 247 28.66 Uridylate kinaseOS = Streptococcus Intracellular pneumoniae (strain CGSP14) GN = pyrH PE= 3 SV = 1 - [B2IP99_STRPS] B2IPF5 10.79 5 834 28.65 Uncharacterizedprotein N- OS = Streptococcus pneumoniae terminally (strain CGSP14) GN =SPCG_0977 anchored PE = 4 SV = 1 - [B2IPF5_STRPS] (No CS) B2IM26 26.1 6318 28.61 Autolysin OS = Streptococcus Intracellular pneumoniae (strainCGSP14) GN = lytA PE = 4 SV = 1 - [B2IM26_STRPS] B2INF6 43.1 5 239 28.52tRNA (guanine-N(1)-)- Intracellular methyltransferase OS = Streptococcuspneumoniae (strain CGSP14) GN = trmD PE = 3 SV = 1 - [TRMD_STRPS] B2ISI015.25 6 833 28.41 Leucine--tRNA ligase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = leuS PE = 3 SV = 1 - [SYL_STRPS] B2IQH820.5 6 439 28.08 Homoserine dehydrogenase Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = hom PE = 3 SV = 1 -[B2IQH8_STRPS] B2INT8 57.89 3 76 27.91 Uncharacterized proteinIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_0763 PE = 4 SV = 1 - [B2INT8_STRPS] B2IMD9 15.64 2 211 27.64 tRNA(guanine-N(7)-)- Intracellular methyltransferase OS = Streptococcuspneumoniae (strain CGSP14) GN = trmB PE = 3 SV = 1 - [TRMB_STRPS] B2IQS226.88 4 253 27.63 SpoU rRNA methylase family Intracellular protein OS =Streptococcus pneumoniae (strain CGSP14) GN = trmH PE = 3 SV = 1 -[B2IQS2_STRPS] B2IR73 16.69 5 623 27.4 ABC transporter, ATP-bindingIntracellular protein OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1540 PE = 4 SV = 1 - [B2IR73_STRPS] B2IPP4 10.12 6 662 27.29 UvrABCsystem protein B Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = uvrB PE = 3 SV = 1 - [UVRB_STRPS] B2IQK6 26.91 4 301 27.27UDP-N- Intracellular acetylenolpyruvoylglucosamine reductase OS =Streptococcus pneumoniae (strain CGSP14) GN = murB PE = 3 SV = 1 -[MURB_STRPS] B2IP98 25.45 6 444 27.26 Methylenetetrahydrofolate--tRNA-Intracellular (uracil-5-)-methyltransferase TrmFO OS = Streptococcuspneumoniae (strain CGSP14) GN = trmFO PE = 3 SV = 2 - [TRMFO_STRPS]B2IQK5 19.48 5 385 27.06 Spermidine/putrescine import ATP- Intracellularbinding protein PotA OS = Streptococcus pneumoniae (strain CGSP14) GN =potA PE = 3 SV = 1 - [B2IQK5_STRPS] B2INN3 30.61 7 392 26.86 Secreted 45kd protein Secretory OS = Streptococcus pneumoniae (released) (strainCGSP14) GN = SPCG_2182 (with PE = 4 SV = 1 - [B2INN3_STRPS] CS) B2IN2040.78 4 103 26.71 Uncharacterized protein Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = SPCG_2067 PE = 4 SV = 1 -[B2IN20_STRPS] B2IPN6 8.14 6 1179 26.7 Chromosome partition protein SmcIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = smc PE= 3 SV = 1 - [B2IPN6_STRPS] B2IRW9 52.07 5 121 26.67 Transcriptionalregulator, GntR Intracellular family OS = Streptococcus pneumoniae(strain CGSP14) GN = SPCG_1686 PE = 4 SV = 1 - [B2IRW9_STRPS] B2IS8413.47 2 438 26.55 Ribosomal RNA small subunit Intracellularmethyltransferase B OS = Streptococcus pneumoniae (strain CGSP14) GN =sunL PE = 3 SV = 1 - [B2IS84_STRPS] B2IMX9 19.88 7 332 26.49Pneumococcal surface protein, Lipid putative OS = Streptococcus anchoredpneumoniae (strain CGSP14) GN = SPCG_0623 PE = 4 SV = 1 - [B2IMX9_STRPS]B2IPI5 8.69 4 541 26.36 RNA methyltransferase, TrmA Intracellular familyOS = Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1007 PE = 3 SV =1 - [B2IPI5_STRPS] B2IPS8 15.14 7 555 25.77 DNA repair protein RecNIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = recN PE= 3 SV = 1 - [B2IPS8_STRPS] B2INY5 17.06 6 551 25.65 DNA polymerase IIIsubunits Intracellular gamma and tau OS = Streptococcus pneumoniae(strain CGSP14) GN = dnaX PE = 4 SV = 1 - [B2INY5_STRPS] B2ISU0 10.1 4594 25.61 ABC transporter, ATP- Multi- binding/permease proteintransmembrane OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1815 PE = 4 SV = 1 - [B2ISU0_STRPS] B2INU6 24.53 6 424 25.32Phosphopentomutase Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = deoB PE = 3 SV = 1 - [B2INU6_STRPS] B2IM77 17.44 2 28125.26 Undecaprenyl-diphosphatase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = uppP PE = 3 SV = 1 - [UPPP_STRPS] B2IS7621.11 5 398 25.26 Hydroxymethylglutaryl-CoA Intracellular synthase OS =Streptococcus pneumoniae (strain CGSP14) GN = mvaS PE = 4 SV = 1 -[B2IS76_STRPS] B2IPD9 20.3 4 266 24.91 Uncharacterized protein N- OS =Streptococcus pneumoniae terminally (strain CGSP14) GN = SPCG_0961anchored PE = 4 SV = 1 - [B2IPD9_STRPS] (No CS) B2IP88 14.52 6 420 24.84Gamma-glutamyl phosphate Intracellular reductase OS = Streptococcuspneumoniae (strain CGSP14) GN = proA PE = 3 SV = 1 - [PROA_STRPS] B2IQU218.36 4 305 24.77 Glycine--tRNA ligase alpha subunit Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = glyQ PE = 3 SV = 1 -[SYGA_STRPS] B2INW0 25 5 220 24.67 Deoxyribose-phosphate aldolaseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = deoC PE= 3 SV = 1 - [DEOC_STRPS] B2IRB6 28.25 6 223 24.63 Cytidylate kinase OS= Streptococcus Intracellular pneumoniae (strain CGSP14) GN = cmk PE = 3SV = 1 - [KCY_STRPS] B2IP93 24.91 5 289 24.4 Ribosomal RNA small subunitIntracellular methyltransferase I OS = Streptococcus pneumoniae (strainCGSP14) GN = rsmI PE = 3 SV = 1 - [B2IP93_STRPS] B2IPN4 29.88 5 32824.28 GMP reductase OS = Streptococcus Intracellular pneumoniae (strainCGSP14) GN = guaC PE = 3 SV = 1 - [GUAC_STRPS] B2INI7 24.9 5 257 24.1Fucose operon repressor, putative Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = fcsR PE = 4 SV = 1 - [B2INI7_STRPS]B2IMP1 31.07 5 206 24.09 Hydrolase, haloacid dehalogenase- Intracellularlike family OS = Streptococcus pneumoniae (strain CGSP14) GN = SPCG_2031PE = 4 SV = 1 - [B2IMP1_STRPS] B2IRV6 17.78 3 343 23.78Phospho-2-dehydro-3- Intracellular deoxyheptonate aldolase OS =Streptococcus pneumoniae (strain CGSP14) GN = aroG PE = 3 SV = 1 -[B2IRV6_STRPS] B2IQ68 27.02 5 359 23.7 Carbamoyl-phosphate synthaseIntracellular small chain OS = Streptococcus pneumoniae (strain CGSP14)GN = carA PE = 3 SV = 1 - [B2IQ68_STRPS] B2IMY1 17.2 3 279 23.45Thymidylate synthase Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = thyA PE = 3 SV = 1 - [B2IMY1_STRPS] B2IM44 17.88 3 27423.39 DNA-entry nuclease N- OS = Streptococcus pneumoniae terminally(strain CGSP14) GN = endA PE = 4 anchored SV = 1 - [B2IM44_STRPS] (NoCS) B2IRI8 22.54 3 213 23.34 Amino acid ABC transporter, ATP-Intracellular binding protein, putative OS = Streptococcus pneumoniae(strain CGSP14) GN = SPCG_0107 PE = 4 SV = 1 - [B2IRI8_STRPS] B2IRQ15.51 2 653 23.17 PTS system IIABC components Multi- OS = Streptococcuspneumoniae transmembrane (strain CGSP14) GN = scrA PE = 4 SV = 1 -[B2IRQ1_STRPS] B2IQR5 19.92 4 246 23.16 CppA protein OS = StreptococcusIntracellular pneumoniae (strain CGSP14) GN = cppA PE = 4 SV = 1 -[B2IQR5_STRPS] B2ILR4 22.22 2 117 23.07 Cell cycle protein GpsBIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = gpsB PE= 3 SV = 1 - [B2ILR4_STRPS] B2IQN2 20.25 4 316 22.86 HPrkinase/phosphorylase Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = hprK PE = 3 SV = 1 - [B2IQN2_STRPS] B2IMA5 25 6 448 22.78Glutamine synthetase, type I Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = glnA PE = 3 SV = 1 - [B2IMA5_STRPS] B2IQ75 33.87 4186 22.38 LemA protein OS = Streptococcus N- pneumoniae (strain CGSP14)terminally GN = lemA PE = 4 SV = 1 - anchored [B2IQ75_STRPS] (No CS)B2INX5 19.41 5 340 21.91 Branched-chain-amino-acid Intracellularaminotransferase OS = Streptococcus pneumoniae (strain CGSP14) GN = ilvEPE = 3 SV = 1 - [B2INX5_STRPS] B2IMY7 43.48 4 207 21.8 Uncharacterizedprotein Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_0630 PE = 4 SV = 1 - [B2IMY7_STRPS] B2IPY6 14.21 4 380 21.63Glucose-1-phosphate Intracellular adenylyltransferase OS = Streptococcuspneumoniae (strain CGSP14) GN = glgC PE = 3 SV = 1 - [GLGC_STRPS] B2IM6920.91 4 416 21.42 L-threonine dehydratase Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = ilvA PE = 3 SV = 1 -[B2IM69_STRPS] B2INA8 35.36 5 263 21.29 Phosphomethylpyrimidine kinaseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = thiD PE= 4 SV = 1 - [B2INA8_STRPS] B2ISK9 13.48 3 267 21.26 PTS system,mannose-specific IIC Multi- component OS = Streptococcus transmembranepneumoniae (strain CGSP14) (Lipid GN = manM PE = 4 SV = 1 - modified[B2ISK9_STRPS] N- termini) B2IQ32 12.93 3 441 21.17 Glycosyltransferase, group 1 Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = SPCG_1204 PE = 4 SV = 1 - [B2IQ32_STRPS] B2IM58 8.12 3 30820.91 Membrane protein insertase YidC Multi- OS = Streptococcuspneumoniae transmembrane (strain CGSP14) GN = yidC PE = 3 SV = 1 -[B2IM58_STRPS] B2INR5 26.54 4 309 20.79 Oxidoreductase, aldo/ketoIntracellular reductase family OS = Streptococcus pneumoniae (strainCGSP14) GN = SPCG_0740 PE = 4 SV = 1 - [B2INR5_STRPS] B2INC9 36.22 3 19620.35 ATP-dependent Clp protease Intracellular proteolytic subunit OS =Streptococcus pneumoniae (strain CGSP14) GN = clpP PE = 3 SV = 1 -[CLPP_STRPS] B2IQ35 25.75 4 369 20.25 RNA polymerase sigma factor SigAIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rpoD PE= 3 SV = 1 - [B2IQ35_STRPS] B2IMR7 20 3 215 20.15 ABC transporter,ATP-binding Intracellular protein Vexp2 OS = Streptococcus pneumoniae(strain CGSP14) GN = SPCG_0561 PE = 4 SV = 1 - [B2IMR7_STRPS]

TABLE 6 Proteins identified in MP from serotype 3. The accession numbersrefer to SwissProt on the date of filing. Accession # Coverage #Peptides #AAs Score Description Localization B2IQV2 90.84 45 404 2064.28Elongation factor Tu Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = tuf PE = 3 SV = 1 - [B2IQV2_STRPS] B2IRT9 69.68 49 4421423.4 Sugar ABC transporter, sugar- Lipid binding protein OS =Streptococcus anchored pneumoniae (strain CGSP14) GN = SPCG_1656 PE = 4SV = 1 - [B2IRT9_STRPS] B2INY1 82.25 60 400 1396.37 30S ribosomalprotein S1 Intracellular OS = Streptococcus pneumoniae (strain CGSP14)GN = rpsA PE = 4 SV = 1 - [B2INY1_STRPS] B2ISJ9 86.15 57 693 1352.29Elongation factor G Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = fusA PE = 3 SV = 1 - [EFG_STRPS] B2IPX8 86.64 41 4341214.93 Enolase OS = Streptococcus Intracellular pneumoniae (strainCGSP14) GN = eno PE = 3 SV = 1 - [ENO_STRPS] B2INB2 75.63 49 591 1080.64Pyruvate oxidase OS = Streptococcus Intracellular pneumoniae (strainCGSP14) GN = spxB PE = 3 SV = 1 - [B2INB2_STRPS] B2IN25 76.44 47 4501068.79 Maltose/maltodextrin ABC Lipid transporter,maltose/maltodextrin- anchored binding protein OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_2072 PE = 4 SV = 1 - [B2IN25_STRPS]B2INK4 76.15 51 608 829.85 Uncharacterized protein N- OS = Streptococcuspneumoniae terminally (strain CGSP14) GN = SPCG_2153 anchored PE = 4 SV= 1 - [B2INK4_STRPS] (No CS) B2ISK8 52.48 17 303 715.49 PTS system,mannose-specific IID Multi- component OS = Streptococcus transmembranepneumoniae (strain CGSP14) GN = manN PE = 4 SV = 1 - [B2ISK8_STRPS]B2IR98 86.7 33 376 633.37 Sugar ABC transporter, ATP-bindingIntracellular protein OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1565 PE = 3 SV = 1 - [B2IR98_STRPS] B2IMI7 75.21 29 359 616.73Glyceraldehyde-3-phosphate Intracellular dehydrogenase OS =Streptococcus pneumoniae (strain CGSP14) GN = gapA PE = 3 SV = 1 -[B2IMI7_STRPS] B2ISL0 55.12 20 332 608.4 PTS system, mannose-specificIIAB Intracellular components OS = Streptococcus pneumoniae (strainCGSP14) GN = manL PE = 4 SV = 1 - [B2ISL0_STRPS] B2IRH0 78.91 39 512573.11 ABC transporter, substrate-binding Lipid protein OS =Streptococcus anchored pneumoniae (strain CGSP14) GN = SPCG_0089 PE = 4SV = 1 - [B2IRH0_STRPS] B2IS43 74.01 23 277 570.24 50S ribosomal proteinL2 Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rplBPE = 3 SV = 1 - [RL2_STRPS] B2INK5 70.12 34 502 534.8 Glycerol kinase OS= Streptococcus Intracellular pneumoniae (strain CGSP14) GN = glpK PE =3 SV = 1 - [GLPK_STRPS] B2IRV7 73.6 55 837 530.96 Protein translocasesubunit SecA Intracellular OS = Streptococcus pneumoniae (strain CGSP14)GN = secA PE = 3 SV = 1 - [B2IRV7_STRPS] B2INW2 65.51 28 374 519.89Lipoprotein OS = Streptococcus Lipid pneumoniae (strain CGSP14) anchoredGN = SPCG_0787 PE = 4 SV = 1 - [B2INW2_STRPS] B2IPD4 58.15 28 313 480.11Foldase protein PrsA Lipid OS = Streptococcus pneumoniae anchored(strain CGSP14) GN = prsA PE = 3 SV = 1 - [PRSA_STRPS] B2IR00 59.97 38652 462.58 ATP-dependent zinc Multi- metalloprotease FtsH transmembraneOS = Streptococcus pneumoniae (strain CGSP14) GN = ftsH PE = 3 SV = 1 -[B2IR00_STRPS] B2IM39 54.12 57 1225 460.95 DNA-directed RNA polymeraseIntracellular subunit beta′ OS = Streptococcus pneumoniae (strainCGSP14) GN = rpoC PE = 3 SV = 1 - [RPOC_STRPS] B2IM81 63.18 37 774432.81 Formate acetyltransferase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = pfl PE = 4 SV = 1 - [B2IM81_STRPS]B2IM49 72.73 25 330 394.5 Aspartate--ammonia ligase Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = asnA PE = 3 SV = 1 -[ASNA_STRPS] B2IN96 69.05 25 378 394.18 Lactate oxidase OS =Streptococcus Intracellular pneumoniae (strain CGSP14) GN = lctO PE = 4SV = 1 - [B2IN96_STRPS] B2IPT8 93.56 27 326 382.46 Tagatose1,6-diphosphate aldolase Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = lacD PE = 3 SV = 1 - [B2IPT8_STRPS] B2INN2 73.36 26259 380.63 30S ribosomal protein S2 Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = rpsB PE = 3 SV = 2 - [RS2_STRPS] B2IQX086.32 28 468 375.02 ATP synthase subunit beta Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = atpD PE = 3 SV = 1 -[ATPB_STRPS] B2IMA2 85.18 25 398 374.17 Phosphoglycerate kinaseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = pgk PE= 3 SV = 1 - [PGK_STRPS] B2IS46 81.11 26 217 368.3 30S ribosomal proteinS3 Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rpsCPE = 3 SV = 1 - [RS3_STRPS] B2ISZ0 64.49 31 659 336.33 Oligopeptide ABCtransporter, Lipid oligopeptide-binding protein AmiA anchored OS =Streptococcus pneumoniae (strain CGSP14) GN = amiA PE = 4 SV = 1 -[B2ISZ0_STRPS] B2ISG3 77.23 21 202 332.38 General stress protein 24,putative Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN= SPCG_1781 PE = 4 SV = 1 - [B2ISG3_STRPS] B2IML8 69.46 25 406 332.33Acetate kinase OS = Streptococcus Intracellular pneumoniae (strainCGSP14) GN = ackA PE = 3 SV = 1 - [B2IML8_STRPS] B2IP54 71.66 27 501328.6 Pyruvate kinase OS = Streptococcus Intracellular pneumoniae(strain CGSP14) GN = SPCG_0873 PE = 3 SV = 1 - [B2IP54_STRPS] B2IN1057.69 12 338 324.09 Glycerol-3-phosphate Intracellular dehydrogenase[NAD(P)+] OS = Streptococcus pneumoniae (strain CGSP14) GN = gpsA PE = 3SV = 1 - [GPDA_STRPS] B2IRR1 66.09 12 230 318.072,3-bisphosphoglycerate- Intracellular dependent phosphoglycerate mutaseOS = Streptococcus pneumoniae (strain CGSP14) GN = gpmA PE = 3 SV = 1 -[GPMA_STRPS] B2IRI6 36.84 5 95 310.42 Bacteriocin, putative N- OS =Streptococcus pneumoniae terminally (strain CGSP14) GN = SPCG_0105anchored PE = 4 SV = 1 - [B2IRI6_STRPS] (with CS) B2IQY1 57.35 26 551302.6 Uncharacterized protein Intracellular/ OS = Streptococcuspneumoniae TMH (strain CGSP14) GN = SPCG_1503 start PE = 4 SV = 1 -[B2IQY1_STRPS] AFTER 60 B2ISJ7 67.88 14 137 301.47 30S ribosomal proteinS12 Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =rpsL PE = 3 SV = 1 - [RS12_STRPS] B2IQX2 38.92 21 501 285.3 ATP synthasesubunit alpha Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = atpA PE = 3 SV = 1 - [ATPA_STRPS] B2IP53 50.15 16 335269.91 ATP-dependent 6- Intracellular phosphofructokinase OS =Streptococcus pneumoniae (strain CGSP14) GN = pfkA PE = 3 SV = 1 -[PFKA_STRPS] B2IR47 51.69 27 652 263.77 Oligopeptide ABC transporter,Lipid oligopeptide-binding protein AliB anchored OS = Streptococcuspneumoniae (strain CGSP14) GN = aliB PE = 4 SV = 1 - [B2IR47_STRPS]B2ISZ6 50.36 17 419 260.6 Sugar ABC transporter, sugar- Lipid bindingprotein OS = Streptococcus anchored pneumoniae (strain CGSP14) GN =SPCG_1871 PE = 4 SV = 1 - [B2ISZ6_STRPS] B2IMB5 52.72 24 607 254.75Chaperone protein DnaK Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = dnaK PE = 3 SV = 1 - [DNAK_STRPS] B2INT1 42.12 26584 253.89 Septation ring formation regulator N- EzrA OS = Streptococcusterminally pneumoniae (strain CGSP14) anchored GN = ezrA PE = 3 SV = 1 -(No CS) [B2INT1_STRPS] B2IM12 66.03 26 471 237.78 Pneumolysin OS =Streptococcus Intracellular pneumoniae (strain CGSP14) GN = ply PE = 4SV = 1 - [B2IM12_STRPS] B2IS47 55.47 11 137 228.66 50S ribosomal proteinL16 Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =rplP PE = 3 SV = 1 - [RL16_STRPS] B2IRM9 68.48 18 276 218.21 ABCtransporter, substrate-binding Lipid protein OS = Streptococcus anchoredpneumoniae (strain CGSP14) GN = SPCG_0152 PE = 4 SV = 1 - [B2IRM9_STRPS]B2IRG4 64.53 23 203 217.88 30S ribosomal protein S4 Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = rpsD PE = 3 SV = 1 -[RS4_STRPS] B2IS57 86.59 19 164 213.33 30S ribosomal protein S5Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rpsE PE= 3 SV = 1 - [B2IS57_STRPS] B2IM40 44.9 29 824 208.41 DNA-directed RNApolymerase Intracellular subunit beta OS = Streptococcus pneumoniae(strain CGSP14) GN = rpoB PE = 3 SV = 1 - [B2IM40_STRPS] B2ISM3 87.16 20148 207.33 50S ribosomal protein L13 Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = rplM PE = 3 SV = 1 - [B2ISM3_STRPS]B2IQ88 76.52 13 115 207.04 50S ribosomal protein L19 Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = rplS PE = 3 SV = 1 -[RL19_STRPS] B2ISD1 80.77 12 104 201.22 Thioredoxin OS = StreptococcusIntracellular pneumoniae (strain CGSP14) GN = trxA PE = 3 SV = 1 -[B2ISD1_STRPS] B2IS40 62.98 15 208 197.25 50S ribosomal protein L3Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rplC PE= 3 SV = 1 - [RL3_STRPS] B2IMJ9 41.57 28 890 195.17 Aldehyde-alcoholdehydrogenase Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = adhE PE = 3 SV = 1 - [B2IMJ9_STRPS] B2IPW4 45.86 24 567191.95 Dihydrolipoyl dehydrogenase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = acoL PE = 4 SV = 1 - [B2IPW4_STRPS]B2IP07 28.31 14 650 190.79 PTS system, fructose specific IIABC Multi-components OS = Streptococcus transmembrane pneumoniae (strain CGSP14)GN = fruA PE = 4 SV = 1 - [B2IP07_STRPS] B2IS55 84.27 13 178 180.96 50Sribosomal protein L6 Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = rplF PE = 3 SV = 1 - [RL6_STRPS] B2IME4 33.44 20 930 180.14Translation initiation factor IF-2 Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = infB PE = 3 SV = 1 - [IF2_STRPS] B2ISW053.72 12 242 178.66 Choline transporter Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_1835 PE = 4 SV = 1 - [B2ISW0_STRPS]B2IS66 43.31 8 127 176.98 30S ribosomal protein S11 Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = rpsK PE = 3 SV = 1 -[RS11_STRPS] B2IPR1 43.98 13 332 173.61 L-lactate dehydrogenaseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = ldh PE= 3 SV = 1 - [B2IPR1_STRPS] B2IRS1 44.15 15 419 169.63 Cell divisionprotein FtsZ Intracellular OS = Streptococcus pneumoniae (strain CGSP14)GN = ftsZ PE = 3 SV = 1 - [B2IRS1_STRPS] B2IPP2 36.89 22 721 166.14Amino acid ABC transporter, amino Multi- acid-binding protein/permeasetransmembrane protein OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1062 PE = 3 SV = 1 - [B2IPP2_STRPS] B2ILZ5 43.89 21 540 165.77 60kDa chaperonin Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = groL PE = 3 SV = 1 - [CH60_STRPS] B2IS41 65.22 16 207162.59 50S ribosomal protein L4 Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = rplD PE = 3 SV = 1 - [RL4_STRPS] B2ILW675.31 15 243 162.56 3-ketoacyl-(Acyl-carrier-protein) Intracellularreductase OS = Streptococcus pneumoniae (strain CGSP14) GN = fabG PE = 4SV = 1 - [B2ILW6_STRPS] B2ISY9 34.54 14 498 161.94 Oligopeptide ABCtransporter, Multi- permease protein AmiC transmembrane OS =Streptococcus pneumoniae (strain CGSP14) GN = amiC PE = 3 SV = 1 -[B2ISY9_STRPS] B2ISY6 58.31 18 331 161.56 Oligopeptide ABC transporter,ATP- Intracellular binding protein AmiF OS = Streptococcus pneumoniae(strain CGSP14) GN = amiF PE = 3 SV = 1 - [B2ISY6_STRPS] B2IS52 77.22 13180 159.07 50S ribosomal protein L5 Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = rplE PE = 3 SV = 1 - [RL5_STRPS] B2ILW750.24 10 414 153.03 3-oxoacyl-[acyl-carrier-protein] Intracellularsynthase 2 OS = Streptococcus pneumoniae (strain CGSP14) GN = fabF PE =3 SV = 1 - [B2ILW7_STRPS] B2ISY7 46.76 16 355 151.21 Oligopeptide ABCtransporter, ATP- Intracellular binding protein AmiE OS = Streptococcuspneumoniae (strain CGSP14) GN = amiE PE = 3 SV = 1 - [B2ISY7_STRPS]B2ILU5 34.89 13 427 150.71 Trigger factor OS = StreptococcusIntracellular pneumoniae (strain CGSP14) GN = tig PE = 3 SV = 1 -[TIG_STRPS] B2IMU8 55.46 15 229 148.33 50S ribosomal protein L1Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rplA PE= 3 SV = 1 - [RL1_STRPS] B2IS44 56.99 10 93 148.01 30S ribosomal proteinS19 Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =rpsS PE = 3 SV = 1 - [RS19_STRPS] B2IQT6 47.49 18 459 146.83 NADHoxidase OS = Streptococcus Intracellular pneumoniae (strain CGSP14) GN =nox PE = 4 SV = 1 - [B2IQT6_STRPS] B2INP5 40.65 15 492 145.57Inosine-5′-monophosphate Intracellular dehydrogenase OS = Streptococcuspneumoniae (strain CGSP14) GN = imdH PE = 3 SV = 1 - [B2INP5_STRPS]B2INF2 51.27 20 513 140.36 ABC transporter, ATP-binding Intracellularprotein OS = Streptococcus pneumoniae (strain CGSP14) GN = SPCG_0719 PE= 4 SV = 1 - [B2INF2_STRPS] B2IRS2 50.33 12 457 140.22 Cell divisionprotein FtsA Intracellular OS = Streptococcus pneumoniae (strain CGSP14)GN = ftsZ PE = 3 SV = 1 - [B2IRS2_STRPS] B2IPY5 51.98 15 379 138.63Glycogen biosynthesis protein GlgD Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = glgD PE = 4 SV = 1 - [B2IPY5_STRPS]B2IQH2 47.59 12 187 137.62 50S ribosomal protein L10 Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = rplJ PE = 3 SV = 1 -[B2IQH2_STRPS] B2IPM7 29.64 14 577 137.31 Phosphoenolpyruvate-proteinIntracellular phosphotransferase OS = Streptococcus pneumoniae (strainCGSP14) GN = SPCG_1121 PE = 3 SV = 1 - [B2IPM7_STRPS] B2ISJ8 57.69 11156 133.5 30S ribosomal protein S7 Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = rpsG PE = 3 SV = 1 - [RS7_STRPS] B2IS5474.24 10 132 133.28 30S ribosomal protein S8 Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = rpsH PE = 3 SV = 1 -[RS8_STRPS] B2INZ1 46.38 17 470 132.61 FeS assembly protein SufBIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_0816 PE = 4 SV = 1 - [B2INZ1_STRPS] B2IS53 62.92 10 89 131.91 30Sribosomal protein S14 Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = rpsN PE = 3 SV = 1 - [RS14_STRPS] B2IS59 63.7 9 146131.45 50S ribosomal protein L15 Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = rplO PE = 3 SV = 1 - [RL15_STRPS] B2INH550.5 17 299 127.85 SPFH domain/Band 7 family N- OS = Streptococcuspneumoniae terminally (strain CGSP14) GN = SPCG_2124 anchored PE = 4 SV= 1 - [B2INH5_STRPS] (No CS) B2IRE3 73.42 12 158 127.54 PTS system, IIBcomponent Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN= SPCG_0062 PE = 4 SV = 1 - [B2IRE3_STRPS] B2IN11 64.88 14 299 125.53UTP--glucose-1-phosphate Secretory uridylyltransferase (released) OS =Streptococcus pneumoniae (with (strain CGSP14) GN = galU PE = 3 SV = 1CS) [B2IN11_STRPS] B2ISI9 43.76 23 617 125.33 Proline--tRNA ligaseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = proS PE= 3 SV = 1 - [SYP_STRPS] B2IPB5 60.5 12 119 125.17 50S ribosomal proteinL20 Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =rplT PE = 3 SV = 1 - [RL20_STRPS] B2IMN9 40.47 11 425 125.01Uncharacterized protein Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = SPCG_2034 PE = 4 SV = 1 - [B2IMN9_STRPS] B2IS9047.86 20 537 123.3 Ribonuclease Y OS = Streptococcus Intracellularpneumoniae (strain CGSP14) GN = rny PE = 3 SV = 1 - [B2IS90_STRPS]B2IPB4 36.36 6 66 122.81 50S ribosomal protein L35 Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = rpmI PE = 3 SV = 1 -[RL35_STRPS] B2IQW2 62.61 5 115 121.34 Bacterocin transport accessoryIntracellular protein OS = Streptococcus pneumoniae (strain CGSP14) GN =bta PE = 4 SV = 1 - [B2IQW2_STRPS] B2INS1 30.9 21 848 120 AminopeptidaseN Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = pepNPE = 4 SV = 1 - [B2INS1_STRPS] B2IPP1 53.66 9 246 116.21 Amino acid ABCtransporter, ATP- Intracellular binding protein OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_1061 PE = 4 SV = 1 - [B2IPP1_STRPS]B2IMN0 34.94 8 352 116.08 Alcohol dehydrogenase, zinc- Intracellularcontaining OS = Streptococcus pneumoniae (strain CGSP14) GN = adh PE = 3SV = 1 - [B2IMN0_STRPS] B2IRN0 53.52 14 284 114.19 Lipoprotein OS =Streptococcus Lipid pneumoniae (strain CGSP14) anchored GN = SPCG_0153PE = 3 SV = 1 - [B2IRN0_STRPS] B2IQZ2 48.66 14 374 110.45Ribosome-binding ATPase YchF Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = ychF PE = 3 SV = 1 - [B2IQZ2_STRPS] B2IQI0 41.57 13344 105.33 Uncharacterized protein Multi- OS = Streptococcus pneumoniaetransmembrane (strain CGSP14) GN = SPCG_1352 PE = 4 SV = 1 -[B2IQI0_STRPS] B2IS49 60.47 7 86 105.08 30S ribosomal protein S17Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rpsQ PE= 3 SV = 1 - [RS17_STRPS] B2IR07 35.97 15 442 104.04 Adenylosuccinatesynthetase Intracellular OS = Streptococcus pneumoniae (strain CGSP14)GN = purA PE = 3 SV = 1 - [B2IR07_STRPS] B2IMU7 42.55 6 141 103.8 50Sribosomal protein L11 Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = rplK PE = 3 SV = 1 - [RL11_STRPS] B2ISV9 21.53 9511 103.52 Choline transporter Multi- OS = Streptococcus pneumoniaetransmembrane (strain CGSP14) GN = SPCG_1834 PE = 3 SV = 1 -[B2ISV9_STRPS] B2IQV9 40.56 16 572 103.11 PhosphoglucomutaseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = pgm PE= 3 SV = 1 - [B2IQV9_STRPS] B2IR17 52.51 12 339 101.9 Ribose-phosphateIntracellular pyrophosphokinase OS = Streptococcus pneumoniae (strainCGSP14) GN = prsA PE = 3 SV = 1 - [B2IR17_STRPS] B2IRQ3 37.38 18 650101.35 Endopeptidase O OS = Streptococcus Intracellular pneumoniae(strain CGSP14) GN = pepO PE = 4 SV = 1 - [B2IRQ3_STRPS] B2IS63 54.72 6212 100 Adenylate kinase OS = Streptococcus Intracellular pneumoniae(strain CGSP14) GN = adk PE = 3 SV = 1 - [KAD_STRPS] B2IMK3 38.15 13 65899.61 Transketolase OS = Streptococcus Intracellular pneumoniae (strainCGSP14) GN = tktA PE = 3 SV = 1 - [B2IMK3_STRPS] B2IMS2 55.97 12 29398.56 Fructose-bisphosphate aldolase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = fba PE = 3 SV = 1 - [B2IMS2_STRPS]B2IMH3 55.78 12 346 96.72 Catabolite control protein A Intracellular OS= Streptococcus pneumoniae (strain CGSP14) GN = ccpA PE = 4 SV = 1 -[B2IMH3_STRPS] B2IPY8 48.31 15 474 96.35 Glyceraldehyde-3-phosphateIntracellular dehydrogenase, NADP-dependent OS = Streptococcuspneumoniae (strain CGSP14) GN = gapN PE = 3 SV = 1 - [B2IPY8_STRPS]B2ILY5 30.81 10 555 96.12 Uncharacterized protein Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = SPCG_0438 PE = 4 SV = 1 -[B2ILY5_STRPS] B2IS42 95.92 11 98 95.91 50S ribosomal protein L23Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rplW PE= 3 SV = 1 - [RL23_STRPS] B2IM28 43.56 13 388 95.69 Protein RecA OS =Streptococcus Intracellular pneumoniae (strain CGSP14) GN = recA PE = 3SV = 1 - [RECA_STRPS] B2IRR6 27.44 10 266 95.52 Cell division proteinDivIVA Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1633 PE = 4 SV = 1 - [B2IRR6_STRPS] B2IS68 64.06 8 128 93.87 50Sribosomal protein L17 Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = rplQ PE = 3 SV = 1 - [RL17_STRPS] B2IPR2 22.47 13841 93.61 DNA gyrase subunit A Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = gyrA PE = 3 SV = 1 - [B2IPR2_STRPS]B2IM73 29.94 12 521 93.46 Amino acid ABC transporter, amino Multi-acid-binding protein/permease transmembrane protein OS = Streptococcuspneumoniae (strain CGSP14) GN = glnH PE = 3 SV = 1 - [B2IM73_STRPS]B2INV5 59.04 7 83 93.23 30S ribosomal protein S20 Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = rpsT PE = 3 SV = 1 -[B2INV5_STRPS] B2IRH9 39.61 21 616 93.09 Capsular polysaccharide Multi-biosynthesis protein, putative transmembrane OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_0098 PE = 4 SV = 1 - [B2IRH9_STRPS]B2INY7 55.13 9 263 92.69 ABC transporter, ATP-binding Intracellularprotein OS = Streptococcus pneumoniae (strain CGSP14) GN = SPCG_0812 PE= 4 SV = 1 - [B2INY7_STRPS] B2INW3 45.21 14 511 92.64 Sugar ABCtransporter, ATP-binding Intracellular protein OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_0788 PE = 4 SV = 1 - [B2INW3_STRPS]B2IQU4 57.5 9 280 89.47 Oxidoreductase, aldo/keto Intracellularreductase family OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1466 PE = 4 SV = 1 - [B2IQU4_STRPS] B2ILR7 36.8 13 481 87.346-phosphogluconate Intracellular dehydrogenase, decarboxylating OS =Streptococcus pneumoniae (strain CGSP14) GN = gnd PE = 3 SV = 1 -[B2ILR7_STRPS] B2IRW4 23.85 14 436 87.08 GTPase Der OS = StreptococcusIntracellular pneumoniae (strain CGSP14) GN = der PE = 3 SV = 1 -[DER_STRPS] B2IQW3 46.76 10 278 86.59 Amino acid ABC transporter, aminoLipid acid-binding protein anchored OS = Streptococcus pneumoniae(strain CGSP14) GN = SPCG_1485 PE = 4 SV = 1 - [B2IQW3_STRPS] B2IP1733.14 8 341 85.78 Iron-compound ABC transporter, Lipid ironcompound-binding protein anchored OS = Streptococcus pneumoniae (strainCGSP14) GN = SPCG_1009 PE = 4 SV = 1 - [B2IPI7_STRPS] B2IS65 58.68 12121 85.63 30S ribosomal protein S13 Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = rpsM PE = 3 SV = 1 - [RS13_STRPS] B2ISM459.23 8 130 85.44 30S ribosomal protein S9 Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = rpsI PE = 3 SV = 1 -[RS9_STRPS] B2INT7 37.9 17 752 85.35 ATP-dependent Clp protease, ATP-Intracellular binding subunit ClpE OS = Streptococcus pneumoniae (strainCGSP14) GN = clpE PE = 3 SV = 1 - [B2INT7_STRPS] B2IMU6 41.18 8 23885.34 Uncharacterized protein Lipid OS = Streptococcus pneumoniaeanchored (strain CGSP14) GN = SPCG_0590 PE = 4 SV = 1 - [B2IMU6_STRPS]B2IQR1 39.42 15 520 84.59 GMP synthase [glutamine- Intracellularhydrolyzing] OS = Streptococcus pneumoniae (strain CGSP14) GN = guaA PE= 3 SV = 1 - [GUAA_STRPS] B2IPT6 68.42 8 171 84.01 Galactose-6-phosphateisomerase Intracellular subunit LacB OS = Streptococcus pneumoniae(strain CGSP14) GN = lacB PE = 3 SV = 1 - [LACB_STRPS] B2INL3 30.25 17810 83.74 ATP-dependent Clp protease, ATP- Intracellular binding subunitOS = Streptococcus pneumoniae (strain CGSP14) GN = SPCG_2162 PE = 3 SV =1 - [B2INL3_STRPS] B2IS51 78.22 10 101 83.43 50S ribosomal protein L24Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rplX PE= 3 SV = 1 - [RL24_STRPS] B2INN1 42.77 10 346 83.3 Elongation factor TsIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = tsf PE= 3 SV = 1 - [EFTS_STRPS] B2IS56 67.18 10 131 82.77 50S ribosomalprotein L18 Intracellular OS = Streptococcus pneumoniae (strain CGSP14)GN = rplR PE = 3 SV = 1 - [B2IS56_STRPS] B2ISX8 27.68 8 336 82.74Non-canonical purine NTP Intracellular pyrophosphatase OS =Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1853 PE = 3 SV = 1 -[B2ISX8_STRPS] B2INT0 24.07 12 648 81.85 DNA gyrase subunit BIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = gyrB PE= 3 SV = 1 - [B2INT0_STRPS] B2IN71 31.13 15 620 81.8 Elongation factorTu family protein Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = SPCG_0638 PE = 4 SV = 1 - [B2IN71_STRPS] B2IPU8 19.19 9 71979.81 Ribonucleoside-diphosphate Intracellular reductase OS =Streptococcus pneumoniae (strain CGSP14) GN = nrdE PE = 3 SV = 1 -[B2IPU8_STRPS] B2INM4 49.45 9 182 79.8 Ribosomal subunit interfaceprotein Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_2173 PE = 4 SV = 1 - [B2INM4_STRPS] B2ISV2 28.57 7 392 79.49Galactokinase OS = Streptococcus Intracellular pneumoniae (strainCGSP14) GN = galK PE = 3 SV = 1 - [B2ISV2_STRPS] B2IMZ0 45.21 9 449 77.7Glucose-6-phosphate isomerase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = pgi PE = 3 SV = 1 - [G6PI_STRPS] B2IRD461.8 6 89 77.46 30S ribosomal protein S15 Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = rpsO PE = 3 SV = 1 -[RS15_STRPS] B2IR65 69.38 11 209 76.49 Uncharacterized proteinIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1532 PE = 4 SV = 1 - [B2IR65_STRPS] B2IR90 43.02 4 172 76.3Non-heme iron-containing ferritin Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_1557 PE = 3 SV = 1 - [B2IR90_STRPS]B2IPY6 28.16 9 380 75.91 Glucose-1-phosphate Intracellularadenylyltransferase OS = Streptococcus pneumoniae (strain CGSP14) GN =glgC PE = 3 SV = 1 - [GLGC_STRPS] B2IN68 38.1 7 126 75.25Uncharacterized protein N- OS = Streptococcus pneumoniae terminally(strain CGSP14) GN = SPCG_0635 anchored PE = 4 SV = 1 - [B2IN68_STRPS](No CS) B2IR60 97.92 8 96 74.99 30S ribosomal protein S6 IntracellularOS = Streptococcus pneumoniae (strain CGSP14) GN = rpsF PE = 3 SV = 1 -[RS6_STRPS] B2IQ02 48.45 7 97 74.43 50S ribosomal protein L27Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rpmA PE= 3 SV = 1 - [RL27_STRPS] B2IM41 40.05 10 377 74.15 Uncharacterizedprotein Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1926 PE = 3 SV = 1 - [B2IM41_STRPS] B2IR71 16.41 12 914 73.99Cation-transporting ATPase, E1-E2 Multi- family OS = Streptococcustransmembrane pneumoniae (strain CGSP14) GN = pacL PE = 3 SV = 1 -[B2IR71_STRPS] B2IQV3 35.99 6 289 72.94 Glycerol uptake facilitatorprotein, Multi- putative OS = Streptococcus transmembrane pneumoniae(strain CGSP14) GN = glpF PE = 3 SV = 1 - [B2IQV3_STRPS] B2ILW5 35.95 9306 72.85 Malonyl CoA-acyl carrier protein Intracellular transacylase OS= Streptococcus pneumoniae (strain CGSP14) GN = fabD PE = 3 SV = 1 -[B2ILW5_STRPS] B2IRQ6 42.81 10 313 71.49 Manganese ABC transporter,Lipid manganese-binding adhesion anchored liprotein OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_1623 PE = 3 SV = 1 - [B2IRQ6_STRPS]B2IQX4 60.98 9 164 71.29 ATP synthase subunit b N- OS = Streptococcuspneumoniae terminally (strain CGSP14) GN = atpF PE = 3 SV = 1 - anchored[ATPF_STRPS] (No CS) B2IPT0 26.59 10 628 71.03 Elongation factor 4Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = lepA PE= 3 SV = 1 - [B2IPT0_STRPS] B2ISJ1 27.74 11 602 69.46Glutamine--fructose-6-phosphate Intracellular aminotransferase[isomerizing] OS = Streptococcus pneumoniae (strain CGSP14) GN = glmS PE= 3 SV = 1 - [B2ISJ1_STRPS] B2IQU1 23.75 13 678 68.47 Glycine--tRNAligase beta subunit Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = glyS PE = 3 SV = 1 - [SYGB_STRPS] B2INH0 41.42 11 338 68.3Ornithine carbamoyltransferase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = arcB PE = 3 SV = 1 - [OTC_STRPS] B2IQL461.42 12 267 68.04 Phosphate import ATP-binding Intracellular proteinPstB OS = Streptococcus pneumoniae (strain CGSP14) GN = pstB PE = 3 SV =1 - [B2IQL4_STRPS] B2ING9 37.9 10 409 65.94 Arginine deiminaseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = arcA PE= 3 SV = 1 - [ARCA_STRPS] B2IR87 27.8 9 410 65.69 ATP-dependent Clpprotease ATP- Intracellular binding subunit CIpX OS = Streptococcuspneumoniae (strain CGSP14) GN = clpX PE = 3 SV = 1 - [CLPX_STRPS] B2IPY419.29 7 477 64.75 Glycogen synthase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = glgA PE = 3 SV = 1 - [GLGA_STRPS] B2ISY036 4 100 63.86 UPF0154 protein SPCG_1855 N- OS = Streptococcuspneumoniae terminally (strain CGSP14) GN = SPCG_1855 anchored PE = 3 SV= 1 - [B2ISY0_STRPS] (No CS) B2IQL1 28.04 6 271 63.33 Amino acid ABCtransporter, amino Lipid acid-binding protein anchored OS =Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1383 PE = 4 SV = 1 -[B2IQL1_STRPS] B2IN60 55 5 60 63.27 50S ribosomal protein L32Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rpmF PE= 3 SV = 1 - [RL32_STRPS] B2IQK0 22.48 15 872 63.2 Alanine--tRNA ligaseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = alaS PE= 3 SV = 1 - [SYA_STRPS] B2INE4 30.43 8 447 62.91 DEAD-box ATP-dependentRNA Intracellular helicase CshB OS = Streptococcus pneumoniae (strainCGSP14) GN = rheB PE = 3 SV = 1 - [B2INE4_STRPS] B2IS50 59.84 8 12262.57 50S ribosomal protein L14 Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = rpIN PE = 3 SV = 1 - [RL14_STRPS] B2IND234.2 8 386 62.51 Branched-chain amino acid ABC Lipid transporter, aminoacid-binding anchored protein OS = Streptococcus pneumoniae (strainCGSP14) GN = SPCG_0699 PE = 4 SV = 1 - [B2IND2_STRPS] B2IRC1 40.12 8 33961.9 UDP-glucose 4-epimerase Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = galE PE = 4 SV = 1 - [B2IRC1_STRPS] B2IR96 77.27 7176 61.33 Adenine phosphoribosyltransferase Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = apt PE = 3 SV = 1 -[B2IR96_STRPS] B2IM46 42.61 9 345 61.11 Uncharacterized protein N- OS =Streptococcus pneumoniae terminally (strain CGSP14) GN = SPCG_1931anchored PE = 4 SV = 1 - [B2IM46_STRPS] (No CS) B2IMG8 28.22 8 404 61Aspartate aminotransferase Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = aspC PE = 4 SV = 1 - [B2IMG8_STRPS] B2IQN0 59.06 5127 60.7 Uncharacterized protein N- OS = Streptococcus pneumoniaeterminally (strain CGSP14) GN = SPCG_1399 anchored PE = 4 SV = 1 -[B2IQN0_STRPS] (No CS) B2IMQ9 22.52 11 737 60.06 PolyribonucleotideIntracellular nucleotidyltransferase OS = Streptococcus pneumoniae(strain CGSP14) GN = pnp PE = 3 SV = 2 - [PNP_STRPS] B2IQ40 24.39 14 89859.69 Phosphoenolpyruvate carboxylase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = ppc PE = 3 SV = 1 - [B2IQ40_STRPS]B2IND9 60.43 10 230 59.42 Cell division ABC transporter, ATP-Intracellular binding protein FtsE OS = Streptococcus pneumoniae (strainCGSP14) GN = ftsE PE = 4 SV = 1 - [B2IND9_STRPS] B2IQU5 27.92 10 48059.4 Peptidoglycan N-acetylglucosamine N- deacetylase A OS =Streptococcus terminally pneumoniae (strain CGSP14) anchored GN = pgdAPE = 4 SV = 1 - (No CS) [B2IQU5_STRPS] B2IQ23 39.06 8 425 59.39Uncharacterized protein Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = SPCG_1195 PE = 4 SV = 1 - [B2IQ23_STRPS] B2INH122.54 6 315 59 Carbamate kinase Secretory OS = Streptococcus pneumoniae(released) (strain CGSP14) GN = arcC PE = 3 SV = 1 - (with[B2INH1_STRPS] CS) B2IRI7 20.76 11 713 58.73 Uncharacterized proteinMulti- OS = Streptococcus pneumoniae transmembrane (strain CGSP14) GN =SPCG_0106 PE = 4 SV = 1 - [B2IRI7_STRPS] B2IMP1 37.86 6 206 56.51Hydrolase, haloacid dehalogenase- Intracellular like family OS =Streptococcus pneumoniae (strain CGSP14) GN = SPCG_2031 PE = 4 SV = 1 -[B2IMP1_STRPS] B2IQH6 45.83 13 312 56.05 Bifunctional methioninesulfoxide N- reductase A/B protein terminally OS = Streptococcuspneumoniae anchored (strain CGSP14) GN = msrA PE = 3 (No CS) SV = 1 -[B2IQH6_STRPS] B2IMB6 28.84 8 378 56.02 Chaperone protein DnaJIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = dnaJ PE= 3 SV = 1 - [B2IMB6_STRPS] B2ISI8 32.46 7 419 55.7 Zinc metalloproteaseMulti- OS = Streptococcus pneumoniae transmembrane (strain CGSP14) GN =eep PE = 3 SV = 1 - [B2ISI8_STRPS] B2IR93 26.64 4 259 55.23Triosephosphate isomerase Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = tpiA PE = 3 SV = 1 - [B2IR93_STRPS] B2ILX0 37.58 11455 54.82 Acetyl-CoA carboxylase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = accC PE = 4 SV = 1 - [B2ILX0_STRPS]B2IS39 65.69 6 102 54.76 30S ribosomal protein S10 Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = rpsJ PE = 3 SV = 1 -[RS10_STRPS] B2IQ29 53.89 9 180 54.16 Hypoxanthine-guanine Intracellularphosphoribosyltransferase OS = Streptococcus pneumoniae (strain CGSP14)GN = hgt PE = 4 SV = 1 - [B2IQZ9_STRPS] B2IMZ6 14.35 10 857 54.16 DNAmismatch repair protein MutS Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = hexA PE = 3 SV = 1 - [B2IMZ6_STRPS] B2IR84 36.49 7296 53.72 Nucleotide-binding protein Intracellular SPCG_1551 OS =Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1551 PE = 3 SV = 1 -[Y1551_STRPS] B2ISX0 31.78 8 321 53.7 Iron-compound ABC transporter,Lipid iron-compound-binding protein anchored OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_1845 PE = 4 SV = 1 - [B2ISX0_STRPS]B2ILX6 56.45 6 186 52.99 Elongation factor P Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = efp PE = 3 SV = 1 -[EFP_STRPS] B2IMZ8 23.45 9 563 52.53 Arginine--tRNA ligase IntracellularOS = Streptococcus pneumoniae (strain CGSP14) GN = argS PE = 3 SV = 1 -[SYR_STRPS] B2INY8 20.48 9 420 52.28 Uncharacterized proteinIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_0813 PE = 4 SV = 1 - [B2INY8_STRPS] B2IRA3 23.52 7 438 52.2Pyridine nucleotide-disulfide Intracellular oxidoreductase OS =Streptococcus pneumoniae (strain CGSP14) GN = merA PE = 3 SV = 1 -[B2IRA3_STRPS] B2IPW2 16.67 6 330 51.71 Acetoin dehydrogenase, E1Intracellular component, beta subunit, putative OS = Streptococcuspneumoniae (strain CGSP14) GN = acoB PE = 4 SV = 1 - [B2IPW2_STRPS]B2IRU1 40.52 9 232 51.05 Putative N-acetylmannosamine-6- Intracellularphosphate 2-epimerase OS = Streptococcus pneumoniae (strain CGSP14) GN =nanE PE = 3 SV = 1 - [B2IRU1_STRPS] B2ISY2 5.67 4 705 50.76 TrehalosePTS system, IIABC Multi- components OS = Streptococcus transmembranepneumoniae (strain CGSP14) GN = treP PE = 4 SV = 1 - [B2ISY2_STRPS]B2IRX3 29.29 10 297 50.56 ABC transporter, ATP-binding Intracellularprotein OS = Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1690 PE= 4 SV = 1 - [B2IRX3_STRPS] B2IQY9 24.72 10 453 50.5 Chromosomalreplication initiator Intracellular protein DnaA OS = Streptococcuspneumoniae (strain CGSP14) GN = dnaA PE = 3 SV = 1 - [DNAA_STRPS] B2IMI136.61 4 183 50.13 Transcription Intracellulartermination/antitermination protein NusG OS = Streptococcus pneumoniae(strain CGSP14) GN = nusG PE = 3 SV = 1 - [B2IMI1_STRPS] B2ING0 66.32 595 49.96 30S ribosomal protein S16 Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = rpsP PE = 3 SV = 1 - [RS16_STRPS] B2INE111.85 6 726 48.69 PTS system, IIABC components Multi- OS = Streptococcuspneumoniae transmembrane (strain CGSP14) GN = ptsG PE = 4 SV = 1 -[B2INE1_STRPS] B2IMN6 33 9 494 48.52 Threonine synthase Intracellular OS= Streptococcus pneumoniae (strain CGSP14) GN = thrC PE = 4 SV = 1 -[B2IMN6_STRPS] B2ILR1 12.8 8 719 47.81 Penicillin-binding protein 1A N-OS = Streptococcus pneumoniae terminally (strain CGSP14) GN = pbp1A PE =4 anchored SV = 1 - [B2ILR1_STRPS] (No CS) B2IR57 22.53 8 466 47.14 Coffamily protein/peptidyl-prolyl Intracellular cis-trans isomerase,cyclophilin type OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1524 PE = 4 SV = 1 - [B2IR57_STRPS] B2IRQ1 10.57 5 653 46.37 PTSsystem IIABC components Multi- OS = Streptococcus pneumoniaetransmembrane (strain CGSP14) GN = scrA PE = 4 SV = 1 - [B2IRQ1_STRPS]B2IRE5 23.91 5 276 46.36 PTS system, IID component Multi- OS =Streptococcus pneumoniae transmembrane (strain CGSP14) GN = SPCG_0064 PE= 4 SV = 1 - [B2IRE5_STRPS] B2IRX0 47.08 8 240 46.21 ABC transporter,ATP-binding Intracellular protein OS = Streptococcus pneumoniae (strainCGSP14) GN = SPCG_1687 PE = 4 SV = 1 - [B2IRX0_STRPS] B2IRI9 40.73 7 30245.81 Amino acid ABC transporter, Lipid periplasmic amino acid-bindinganchored protein, putative OS = Streptococcus pneumoniae (strain CGSP14)GN = SPCG_0108 PE = 4 SV = 1 - [B2IRI9_STRPS] B2IQL3 29.76 6 252 45.8Phosphate import ATP-binding Intracellular protein PstB OS =Streptococcus pneumoniae (strain CGSP14) GN = pstB PE = 3 SV = 1 -[B2IQL3_STRPS] B2IQ98 60.64 5 94 45.67 50S ribosomal protein L31 type BIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rpmE PE= 3 SV = 1 - [B2IQ98_STRPS] B2IRE7 15.8 6 386 44.99 Sugar isomerasedomain protein Intracellular AgaS OS = Streptococcus pneumoniae (strainCGSP14) GN = agaS PE = 4 SV = 1 - [B2IRE7_STRPS] B2IMZ5 21.99 9 56444.92 ABC transporter, ATP- Multi- binding/permease proteintransmembrane OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_2042 PE = 4 SV = 1 - [B2IMZ5_STRPS] B2IRQ7 56.4 7 172 44.31Probable thiol peroxidase Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = tpx PE = 3 SV = 1 - [B2IRQ7_STRPS] B2ILX8 33.61 8488 43.22 Glutamyl-tRNA(Gln) Intracellular amidotransferase subunit A OS= Streptococcus pneumoniae (strain CGSP14) GN = gatA PE = 3 SV = 1 -[GATA_STRPS] B2IM45 29.04 8 427 43.07 UDP-N-acetylglucosamine 1-Intracellular carboxyvinyltransferase OS = Streptococcus pneumoniae(strain CGSP14) GN = murA PE = 3 SV = 1 - [B2IM45_STRPS] B2IM60 25 6 41643.06 Diaminopimelate decarboxylase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = lysA PE = 3 SV = 1 - [B2IM60_STRPS]B2IRA1 21.95 9 524 42.94 DEAD-box ATP-dependent RNA Intracellularhelicase CshA OS = Streptococcus pneumoniae (strain CGSP14) GN = cshA PE= 3 SV = 1 - [B2IRA1_STRPS] B2IR78 17.33 6 450 42.72 Phosphoglucosaminemutase Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =glmM PE = 3 SV = 1 - [GLMM_STRPS] B2IST7 22.06 7 417 42.6 Capsularpolysaccharide Intracellular biosynthesis protein, putative OS =Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1812 PE = 3 SV = 1 -[B2IST7_STRPS] B2INT8 57.89 2 76 41.48 Uncharacterized proteinIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_0763 PE = 4 SV = 1 - [B2INT8_STRPS] B2IQ61 25.95 6 289 41.38Choline kinase OS = Streptococcus Intracellular pneumoniae (strainCGSP14) GN = pck PE = 4 SV = 1 - [B2IQ61_STRPS] B2ILR6 14.66 5 464 41.32Mid-cell-anchored protein Z Multi- OS = Streptococcus pneumoniaetransmembrane (strain CGSP14) GN = mapZ PE = 3 SV = 1 - [B2ILR6_STRPS]B2ILR4 29.91 3 117 40.9 Cell cycle protein GpsB Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = gpsB PE = 3 SV = 1 -[B2ILR4_STRPS] B2IQ25 28.88 8 419 40.5 UDP-N-acetylglucosamine 1-Intracellular carboxyvinyltransferase OS = Streptococcus pneumoniae(strain CGSP14) GN = murZ PE = 3 SV = 1 - [B2IQ25_STRPS] B2IQH8 27.11 8439 40.48 Homoserine dehydrogenase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = hom PE = 3 SV = 1 - [B2IQH8_STRPS]B2IN58 28.96 6 335 40.08 Uncharacterized protein Multi- OS =Streptococcus pneumoniae transmembrane (strain CGSP14) GN = SPCG_2101 PE= 4 SV = 1 - [B2IN58_STRPS] B2IRK5 20.21 7 559 40.01 Ribonuclease J OS =Streptococcus Intracellular pneumoniae (strain CGSP14) GN = rnj PE = 3SV = 1 - [B2IRK5_STRPS] B2INR2 26.51 10 679 39.94 Methionine--tRNAligase Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =metG PE = 3 SV = 1 - [B2INR2_STRPS] B2IQ29 30.5 8 436 39.93 GTPase ObgOS = Streptococcus Intracellular pneumoniae (strain CGSP14) GN = obg PE= 3 SV = 1 - [OBG_STRPS] B2IMY2 73.21 7 56 39.82 Uncharacterized proteinSecretory OS = Streptococcus pneumoniae (released) (strain CGSP14) GN =SPCG_0626 (with PE = 4 SV = 1 - [B2IMY2_STRPS] CS) B2IPW3 33.6 8 37539.59 Dihydrolipoamide acetyltransferase Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = acoC PE = 3 SV = 1 -[B2IPW3_STRPS] B2IRI8 38.5 6 213 39.57 Amino acid ABC transporter, ATP-Intracellular binding protein, putative OS = Streptococcus pneumoniae(strain CGSP14) GN = SPCG_0107 PE = 4 SV = 1 - [B2IRI8_STRPS] B2ISR013.08 6 535 39.53 Glucan 1,6-alpha-glucosidase Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = dexB PE = 4 SV = 1 -[B2ISR0_STRPS] B2IM72 56.91 6 246 39.3 Amino acid ABC transporter, ATP-Intracellular binding protein OS = Streptococcus pneumoniae (strainCGSP14) GN = glnQ PE = 4 SV = 1 - [B2IM72_STRPS] B2IS45 48.25 3 11439.27 50S ribosomal protein L22 Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = rplV PE = 3 SV = 1 - [RL22_STRPS] B2IM2626.1 5 318 39.16 Autolysin OS = Streptococcus Intracellular pneumoniae(strain CGSP14) GN = lytA PE = 4 SV = 1 - [B2IM26_STRPS] B2IM71 46.48 471 38.4 Uncharacterized protein Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_0446 PE = 4 SV = 1 - [B2IM71_STRPS]B2INE0 30.09 8 329 38.01 Cell division protein FtsX Multi- OS =Streptococcus pneumoniae transmembrane (strain CGSP14) GN = ftsX PE = 3SV = 1 - [B2INE0_STRPS] B2IQ19 15.99 8 763 37.52 DNA helicase OS =Streptococcus Intracellular pneumoniae (strain CGSP14) GN = pcrA PE = 3SV = 1 - [B2IQ19_STRPS] B2IRN7 21.52 6 660 37.25 Threonine--tRNA ligaseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = thrS PE= 3 SV = 1 - [B2IRN7_STRPS] B2IS67 24.44 7 311 36.89 DNA-directed RNApolymerase Intracellular subunit alpha OS = Streptococcus pneumoniae(strain CGSP14) GN = rpoA PE = 3 SV = 1 - [B2IS67_STRPS] B2IQA4 21.65 8448 36.16 Glutamate dehydrogenase Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = gdhA PE = 3 SV = 1 - [B2IQA4_STRPS]B2INE5 20.96 6 396 36.08 S-adenosylmethionine synthase Intracellular OS= Streptococcus pneumoniae (strain CGSP14) GN = metK PE = 3 SV = 1 -[METK_STRPS] B2IQN3 51.52 4 66 35.91 30S ribosomal protein S21Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = rpsU PE= 3 SV = 1 - [B2IQN3_STRPS] B2IRK4 18.21 8 637 35.62 tRNA uridine 5-Intracellular carboxymethylaminomethyl modification enzyme MnmG OS =Streptococcus pneumoniae (strain CGSP14) GN = mnmG PE = 3 SV = 1 -[B2IRK4_STRPS] B2IR76 14.89 4 282 35.33 DegV family proteinIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1543 PE = 4 SV = 1 - [B2IR76_STRPS] B2IRT0 25.85 4 294 35.15 ROKfamily protein Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = SPCG_1647 PE = 4 SV = 1 - [B2IRT0_STRPS] B2IR21 58.62 7 14535.06 Uncharacterized protein Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_0033 PE = 4 SV = 1 - [B2IR21_STRPS]B2IPF4 25.4 8 311 35.02 Adhesion lipoprotein Lipid OS = Streptococcuspneumoniae anchored (strain CGSP14) GN = lmb PE = 3 SV = 1 -[B2IPF4_STRPS] B2ILV9 26.21 6 454 34.9 Aspartokinase OS = StreptococcusIntracellular pneumoniae (strain CGSP14) GN = lysC PE = 3 SV = 1 -[B2ILV9_STRPS] B2IPP0 17.37 7 495 34.88 Glucose-6-phosphate 1-Intracellular dehydrogenase OS = Streptococcus pneumoniae (strainCGSP14) GN = zwf PE = 3 SV = 1 - [B2IPP0_STRPS] B2IRU0 8.43 5 510 34.78PTS system, IIBC components Multi- OS = Streptococcus pneumoniaetransmembrane (strain CGSP14) GN = SPCG_1657 PE = 4 SV = 1 -[B2IRU0_STRPS] B2IQX1 34.93 7 292 34.34 ATP synthase gamma chainIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = atpG PE= 3 SV = 1 - [ATPG_STRPS] B2ISZ2 16.94 5 490 34.05 Sucrose phosphorylaseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1867 PE = 4 SV = 1 - [B2ISZ2_STRPS] B2IP00 28.21 6 397 33.73 Serineprotease OS = Streptococcus N- pneumoniae (strain CGSP14) terminally GN= sphtra PE = 4 SV = 1 - anchored [B2IP00_STRPS] (No CS) B2INW0 31.82 5220 33.64 Deoxyribose-phosphate aldolase Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = deoC PE = 3 SV = 1 -[DEOC_STRPS] B2IM98 15.7 6 535 33.16 CTP synthase OS = StreptococcusIntracellular pneumoniae (strain CGSP14) GN = pyrG PE = 3 SV = 1 -[B2IM98_STRPS] B2IMU0 11.8 4 466 33.1 Dipeptidase OS = StreptococcusIntracellular pneumoniae (strain CGSP14) GN = pepV PE = 4 SV = 1 -[B2IMU0_STRPS] B2IPU4 37.15 5 253 33.02 Lactose phosphotransferasesystem Intracellular repressor OS = Streptococcus pneumoniae (strainCGSP14) GN = lacR PE = 4 SV = 1 - [B2IPU4_STRPS] B2IRR9 38.55 5 17932.57 Cell division protein SepF Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = sepF PE = 3 SV = 1 - [SEPF_STRPS] B2IQJ527.56 5 225 32.43 3-dehydroquinate dehydratase Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = aroD PE = 3 SV = 1 -[AROD_STRPS] B2INC8 38.43 7 216 32.37 Uracil phosphoribosyltransferaseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = upp PE= 3 SV = 1 - [B2INC8_STRPS] B2INU6 33.73 8 424 32.09 PhosphopentomutaseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = deoB PE= 3 SV = 1 - [B2INU6_STRPS] B2IME1 24.87 6 378 32.06 TranscriptionIntracellular termination/antitermination protein NusA OS =Streptococcus pneumoniae (strain CGSP14) GN = nusA PE = 3 SV = 1 -[B2IME1_STRPS] B2INN5 22.79 4 272 31.93 Cell shape-determining proteinN- MreC OS = Streptococcus terminally pneumoniae (strain CGSP14)anchored GN = mreC PE = 3 SV = 1 - (No CS) [B2INN5_STRPS] B2IN76 12.22 6450 31.55 UDP-N-acetylmuramoylalanine--D- Intracellular glutamate ligaseOS = Streptococcus pneumoniae (strain CGSP14) GN = murD PE = 3 SV = 1 -[MURD_STRPS] B2INF3 16.1 4 267 31.17 Peptidyl-prolyl cis-trans isomeraseLipid OS = Streptococcus pneumoniae anchored (strain CGSP14) GN = ppiAPE = 3 SV = 1 - [B2INF3_STRPS] B2IQ22 31.82 6 286 30.97 Methionineaminopeptidase Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = map PE = 1 SV = 1 - [B2IQ22_STRPS] B2IS48 70.59 5 68 30.850S ribosomal protein L29 Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = rpmC PE = 3 SV = 1 - [RL29_STRPS] B2IS10 19.57 6649 30.36 DNA mismatch repair protein MutL Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = mutL PE = 3 SV = 1 -[MUTL_STRPS] B2IM44 29.93 6 274 30.16 DNA-entry nuclease N- OS =Streptococcus pneumoniae terminally (strain CGSP14) GN = endA PE = 4anchored SV = 1 - [B2IM44_STRPS] (No CS) B2INF7 54.46 5 112 29.66 ATPcone domain-containing Intracellular protein OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_0728 PE = 4 SV = 1 - [B2INF7_STRPS]B2IS34 13.3 7 737 29.66 Anaerobic ribonucleoside Intracellulartriphosphate reductase OS = Streptococcus pneumoniae (strain CGSP14) GN= SPCG_0212 PE = 4 SV = 1 - [B2IS34_STRPS] B2IN20 40.78 3 103 29.58Uncharacterized protein Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = SPCG_2067 PE = 4 SV = 1 - [B2IN20_STRPS] B2IMB738.95 4 95 29.51 Uncharacterized protein Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = SPCG_0492 PE = 4 SV = 1 -[B2IMB7_STRPS] B2IMY9 20.58 6 486 29.3 Glutamate--tRNA ligaseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = gltX PE= 3 SV = 1 - [SYE_STRPS] B2IQ75 46.77 5 186 29.27 LemA protein OS =Streptococcus N- pneumoniae (strain CGSP14) terminally GN = lemA PE = 4SV = 1 - anchored [B2IQ75_STRPS] (No CS) B2IPN9 20.75 6 429 29.19 Signalrecognition particle receptor Intracellular FtsY OS = Streptococcuspneumoniae (strain CGSP14) GN = ftsY PE = 3 SV = 1 - [B2IPN9_STRPS]B2INM2 45.33 7 150 28.94 50S ribosomal protein L9 Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = rplI PE = 3 SV = 1 -[RL9_STRPS] B2IR80 20.89 5 292 28.84 Uncharacterized protein Multi- OS =Streptococcus pneumoniae transmembrane (strain CGSP14) GN = SPCG_1547 PE= 4 SV = 1 - [B2IR80_STRPS] B2IPW9 32.16 8 283 28.64 Ribosome biogenesisGTPase A Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN= SPCG_1141 PE = 3 SV = 1 - [B2IPW9_STRPS] B2IN61 75.51 4 49 28.27 50Sribosomal protein L33 Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = rpmG PE = 3 SV = 1 - [RL33_STRPS] B2IM15 25.95 5158 28.26 Uncharacterized protein Multi- OS = Streptococcus pneumoniaetransmembrane (strain CGSP14) GN = SPCG_1901 PE = 4 SV = 1 -[B2IM15_STRPS] B2IPZ5 13.9 7 633 28.19 ABC transporter, ATP-bindingIntracellular protein OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1167 PE = 4 SV = 1 - [B2IPZ5_STRPS] B2IMA9 20.94 6 487 28.12 Type Irestriction-modification Intracellular system, M subunit OS =Streptococcus pneumoniae (strain CGSP14) GN = hsdM PE = 4 SV = 1 -[B2IMA9_STRPS] B2IQ67 11.63 7 1058 28.04 Carbamoyl-phosphate synthaseIntracellular large chain OS = Streptococcus pneumoniae (strain CGSP14)GN = carB PE = 3 SV = 1 - [CARB_STRPS] B2IMV8 16.5 4 491 28.03 PTSsystem, IIC component, Multi- putative OS = Streptococcus transmembranepneumoniae (strain CGSP14) GN = SPCG_0602 PE = 4 SV = 1 - [B2IMV8_STRPS]B2INP6 15.54 4 341 28 Tryptophan--tRNA ligase Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = trpS PE = 3 SV = 1 -[B2INP6_STRPS] B2INU9 26.02 4 269 27.83 Purine nucleoside phosphorylaseIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = pnp PE= 3 SV = 1 - [B2INU9_STRPS] B2IMS9 20.8 7 553 27.26 Ribonuclease J OS =Streptococcus Intracellular pneumoniae (strain CGSP14) GN = rnj PE = 3SV = 1 - [B2IMS9_STRPS] B2ILY3 41.94 4 62 27.17 50S ribosomal proteinL28 Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =rpmB PE = 3 SV = 1 - [RL28_STRPS] B2IRW8 34.46 5 177 26.92Transcriptional repressor NrdR Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = nrdR PE = 3 SV = 1 - [B2IRW8_STRPS]B2IMH1 20.97 7 472 26.92 Cof family protein Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1962 PE = 4 SV = 1 -[B2IMH1_STRPS] B2IPM6 13.79 2 87 26.87 Phosphocarrier protein HPrIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1120 PE = 4 SV = 1 - [B2IPM6_STRPS] B2IRB7 27.22 2 158 26.84Uncharacterized protein N- OS = Streptococcus pneumoniae terminally(strain CGSP14) GN = SPCG_1584 anchored PE = 4 SV = 1 - [B2IRB7_STRPS](No CS) B2ILW4 23.46 5 324 26.69 Enoyl-(Acyl-carrier-protein)Intracellular reductase OS = Streptococcus pneumoniae (strain CGSP14) GN= fabK PE = 4 SV = 1 - [B2ILW4_STRPS] B2IN41 16.32 7 429 26.52Histidine--tRNA ligase Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = hisS PE = 3 SV = 1 - [SYH_STRPS] B2IQU2 16.07 3 30526.2 Glycine--tRNA ligase alpha subunit Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = glyQ PE = 3 SV = 1 - [SYGA_STRPS] B2IPW123.91 4 322 26.19 Acetoin dehydrogenase, E1 Intracellular component,alpha subunit, putative OS = Streptococcus pneumoniae (strain CGSP14) GN= acoA PE = 4 SV = 1 - [B2IPW1_STRPS] B2ISG1 25.79 5 190 26.04Uncharacterized protein Multi- OS = Streptococcus pneumoniaetransmembrane (strain CGSP14) GN = SPCG_1779 PE = 4 SV = 1 -[B2ISG1_STRPS] B2IPZ7 19.35 4 279 25.42 DegV family proteinIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =SPCG_1169 PE = 4 SV = 1 - [B2IPZ7_STRPS] B2ISK9 13.48 4 267 25.25 PTSsystem, mannose-specific IIC Multi- component OS = Streptococcustransmembrane pneumoniae (strain CGSP14) (Lipid GN = manM PE = 4 SV =1 - modified [B2ISK9_STRPS] N- termini) B2IS70 20.32 5 433 25.16 UPF0210protein SPCG_0246 Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = SPCG_0246 PE = 3 SV = 1 - [B2IS70_STRPS] B2INL6 15.22 4 33524.97 ABC transporter, substrate-binding Lipid protein, putative OS =Streptococcus anchored pneumoniae (strain CGSP14) GN = SPCG_2165 PE = 4SV = 1 - [B2INL6_STRPS] B2IPF2 46.6 4 191 24.92 Thioredoxin familyprotein Lipid OS = Streptococcus pneumoniae anchored (strain CGSP14) GN= SPCG_0974 PE = 4 SV = 1 - [B2IPF2_STRPS] B2INJ2 24.59 7 427 24.82 DltDprotein OS = Streptococcus N- pneumoniae (strain CGSP14) terminally GN =dltD PE = 4 SV = 1 - anchored [B2INJ2_STRPS] (No CS) B2IPH6 13.65 3 35924.36 Peptide chain release factor 1 Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = prfA PE = 3 SV = 1 - [RF1_STRPS] B2ILY020.43 4 514 24.35 Peptide chain release factor 3 Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = prfC PE = 3 SV = 1 -[RF3_STRPS] B2ILV5 20.88 4 182 24.04 Alkyl hydroperoxide reductaseIntracellular AhpD OS = Streptococcus pneumoniae (strain CGSP14) GN =mip PE = 3 SV = 1 - [B2ILV5_STRPS] B2IRX2 21.8 4 399 23.82Uncharacterized protein Multi- OS = Streptococcus pneumoniaetransmembrane (strain CGSP14) GN = SPCG_1689 PE = 4 SV = 1 -[B2IRX2_STRPS] B2IM58 11.36 4 308 23.79 Membrane protein insertase YidCMulti- OS = Streptococcus pneumoniae transmembrane (strain CGSP14) GN =yidC PE = 3 SV = 1 - [B2IM58_STRPS] B2IR27 23.03 4 330 23.79 Phosphateacyltransferase Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = plsX PE = 3 SV = 1 - [PLSX_STRPS] B2IQL7 22.95 4 292 23.74Phosphate ABC transporter, Lipid phosphate-binding protein, anchoredputative OS = Streptococcus pneumoniae (strain CGSP14) GN = SPCG_1389 PE= 4 SV = 1 - [B2IQL7_STRPS] B2IR86 41.62 7 197 23.54 ProbableGTP-binding protein EngB Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = engB PE = 3 SV = 1 - [ENGB_STRPS] B2IP77 21 5 41923.53 Uncharacterized protein Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_0896 PE = 4 SV = 1 - [B2IP77_STRPS]B2ISL2 23.6 5 339 23.5 Alcohol dehydrogenase Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = adhP PE = 3 SV = 1 -[B2ISL2_STRPS] B2IPR3 21.05 5 247 23.35 Uncharacterized protein N- OS =Streptococcus pneumoniae terminally (strain CGSP14) GN = SPCG_1083anchored PE = 4 SV = 1 - [B2IPR3_STRPS] (with CS) B2IR97 25 6 252 23.12Methyltransferase, putative Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = SPCG_1564 PE = 4 SV = 1 - [B2IR97_STRPS] B2IS9824.4 4 209 22.81 Probable nicotinate-nucleotide Intracellularadenylyltransferase OS = Streptococcus pneumoniae (strain CGSP14) GN =nadD PE = 3 SV = 1 - [B2IS98_STRPS] B2IQS3 22.11 4 303 22.58 Thioredoxinreductase Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN= trxB PE = 3 SV = 1 - [B2IQS3_STRPS] B2IPW5 31.61 6 329 22.56Lipoate-protein ligase, putative Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_1137 PE = 4 SV = 1 - [B2IPW5_STRPS]B2IMB4 30.77 5 182 22.24 Protein GrpE OS = Streptococcus Intracellularpneumoniae (strain CGSP14) GN = grpE PE = 3 SV = 1 - [B2IMB4_STRPS]B2ILY4 43.8 3 121 22.23 Uncharacterized protein Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = asp23 PE = 4 SV = 1 -[B2ILY4_STRPS] B2IPH3 18.37 7 479 22.18 tRNA modification GTPase MnmEIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = thdF PE= 3 SV = 1 - [B2IPH3_STRPS] B2INP7 17.59 6 540 22.16 ABC transporter,ATP-binding Intracellular protein OS = Streptococcus pneumoniae (strainCGSP14) GN = SPCG_2196 PE = 4 SV = 1 - [B2INP7_STRPS] B2IP74 14.66 5 49121.9 Lysine decarboxylase Intracellular OS = Streptococcus pneumoniae(strain CGSP14) GN = cad PE = 4 SV = 1 - [B2IP74_STRPS] B2ISG4 52.24 267 21.88 Uncharacterized protein Intracellular OS = Streptococcuspneumoniae (strain CGSP14) GN = SPCG_1782 PE = 3 SV = 1 - [B2ISG4_STRPS]B2ISY1 17.74 7 541 21.79 Dextran glucosidase DexS, putativeIntracellular OS = Streptococcus pneumoniae (strain CGSP14) GN = dexS PE= 4 SV = 1 - [B2ISY1_STRPS] B2IMA5 15.4 5 448 21.67 Glutaminesynthetase, type I Intracellular OS = Streptococcus pneumoniae (strainCGSP14) GN = glnA PE = 3 SV = 1 - [B2IMA5_STRPS] B2IN77 24.72 6 35221.61 UDP-N-acetylglucosamine--N- Intracellularacetylmuramyl-(pentapeptide) pyrophosphoryl-undecaprenol N-acetylglucosamine transferase OS = Streptococcus pneumoniae (strainCGSP14) GN = murG PE = 3 SV = 1 - [MURG_STRPS] B2IQ12 25.39 5 319 21.48Ribose-phosphate Intracellular pyrophosphokinase OS = Streptococcuspneumoniae (strain CGSP14) GN = prs PE = 3 SV = 1 - [B2IQ12_STRPS]B2ISL4 16.3 4 270 21.32 Cof family protein Intracellular OS =Streptococcus pneumoniae (strain CGSP14) GN = SPCG_0299 PE = 4 SV = 1 -[B2ISL4_STRPS] B2IQ78 13.38 5 523 21.2 Signal recognition particleprotein Intracellular OS = Streptococcus pneumoniae (strain CGSP14) GN =ffh PE = 3 SV = 1 - [B2IQ78_STRPS] B2IS71 21.69 5 249 21.06Phosphoglycerate mutase family Intracellular protein OS = Streptococcuspneumoniae (strain CGSP14) GN = gpmB PE = 4 SV = 1 - [B2IS71_STRPS]B2IM30 15.38 5 338 20.64 Transcriptional regulator, putative SecretoryOS = Streptococcus pneumoniae (released) (strain CGSP14) GN = lytR PE =4 SV = 1 - (with [B2IM30_STRPS] CS) B2IQN9 10.91 4 486 20.01 NicotinateIntracellular phosphoribosyltransferase OS = Streptococcus pneumoniae(strain CGSP14) GN = SPCG_1408 PE = 3 SV = 1 - [B2IQN9_STRPS]

Example 2: Microparticles (MPs) Confer Serotype-Independent ProtectiveImmunity

For each immunization experiment male C57BL/6 wild-type mice ca 5 weeksold were used. Before immunization, mice were anesthetized by inhalationof isofluorane (Abbott) and then intranasally administered with 50μl/mouse of microparticles (MP) combined with the adjuvant aluminiumhydroxide (Sigma Aldrich, 10 mg/ml in PBS) or the adjuvant alone for thecontrol groups. Immunization was repeated again after two weeks from thefirst immunization following the same conditions described above. After4 weeks of immunization, mice were infected by intranasal administrationof 50 μl/mouse of 5×10⁶ CFU, for S. pneumoniae type 1 infectionexperiments (FIG. 7 ), or 10⁶ CFU, for S. pneumoniae type 3 infectionexperiments (FIG. 11 ).

All mice were anesthetized by inhalation of isofluorane prior tochallenge with bacteria. After the infection, clinical symptoms of themice were monitored multiple times per day (in accordance with theethical permit). Blood samples (5 μl/mouse) were taken every day ofinfection and level of bacteremia was assessed by plating serialdilutions of blood samples onto blood-agar plates. Mice that reachedhumane end-points were anesthetized by inhalation of isofluorane andintranasal administered with 100 μl/mouse of the fluorescent markerBacterisense 645 (Perkin Elmer) 30 minutes before the sacrifice. Priorto sacrifice, mice were anesthetized again (isofluorane inhalation).After sacrifice, mice were placed in the IVIS Spectrum Imaging System todetect the fluorescent signal of the bacterial infection. After the IVISimaging, lungs and spleens were collected for further analysis.Bacterial amount in the lungs was assessed by CFU count of bacteriaafter plating serial dilutions of lung homogenates onto blood-agarplates.

Intranasal immunization with microparticles (MP) from a serotype 4 S.pneumoniae increased survival after a challenge with serotype 1 S.pneumoniae (FIG. 7A). The increase in survival was significant also incomparison to immunization with MV_(L). The immunization greatly reducedthe bacterial load in the lungs of the mice surviving the challenge withserotype 1 S. pneumoniae (FIG. 7B). Furthermore, mice immunized withserotype 3 MP and then challenged with serotype 3 bacteria showed 100%protection in the pneumonia model, higher than immunization with thecurrently available vaccine PCV13, which only conferred a protection of75% in the model. Moreover, protection against pneumococcal infectionwas absent in mice lacking B cells, suggesting the involvement of anadaptive immune response with anti-pneumococcal antibodies (FIG. 11 ).

Example 3: Serotype-Independent Binding and Opsonophagocitic Activity ofAntibodies Raised Against Pneumococcal Membrane Particles

Antibodies raised against the inventive MP from serotype 4 as antigenwere reactive against Streptococcus pneumoniae of other serotypes, suchas 1, 6B and 3 (FIG. 9 ). In other words, the MP were able to elicitproduction of serotype-independent antibodies, much more so than MV_(L)(FIG. 9B).

The production of anti-MP antibodies in mice was more efficient (inparticular at time point 2 weeks) when the antigen was administeredtogether with an adjuvant. However, satisfactory results were obtainableeven without an adjuvant (FIG. 8 ). Moreover, antibodies in sera wereable to bind both the encapsulated T4 (TIGR4) and the unencapsulatedisogenic mutant T4R to the same extent, indicating that the antibodyresponse was capsule-independent.

Importantly, the anti-MP antibodies showed opsonophagocitic activity,increasing adhesion of bacteria and killing of internalized bacteria byRAW murine macrophages (FIG. 10 ).

Materials and Methods

Bacterial strains and growth conditions. In this study six strains ofStreptococcus pneumoniae were used: TIGR4 or T4 (serotype 4) (22), itsisogenic mutants lacking the cytotoxin pneumolysin (T4Δply) (23), theautolysin LytA (T4ΔlytA) (3), or the capsule (T4R) (24), ATCC 6301(serotype 1) and 1-33 (serotype 3). Bacteria were grown in C+Y medium pH7.9-8.0 (for serotype 4) or THY medium (for serotype 1 and 3) at 37° C.A spectrophotometer (Genesys 20, Thermo Spectronic) was used to followthe growth by monitoring the optical density (OD) at 600 nm.

Isolation and purification of membrane vesicles (MV_(L)) andmicroparticles (MP). For isolation of MV_(L) from a liquid culture,pneumococcal strains (TIGR4, T4Δply or T4ΔlytA) were grown at 37° C. inC+Y medium, pH 7.9-8.0 until OD_(600 nm)=0.9. The culture was thencentrifuged (17,000×g for 30 minutes at 4° C.) to remove bacterial cellsfrom the supernatant. The cell-free supernatant was filtered through a0.22 μm filter (Sarstedt) and centrifuged (120,000×g for 2 hours at 4°C.) to sediment the vesicles. Pellets were washed twice inphosphate-buffered saline (PBS) and resuspended in PBS.

For isolation of microparticles (MP) from bacteria grown on plates, thepneumococcal strains were streaked on blood agar plates and incubatedovernight at 37° C. with 5% CO₂. Bacteria were harvested from plates,resuspended in PBS and MP were pelleted following the same procedure asmentioned above.

Crude MV_(L)/MP preparations were further purified by density gradientcentrifugation using Optiprep™ Density Gradient Medium (Sigma). Pelletedparticle fractions were adjusted to 50% (w/v) Optiprep™ in a totalvolume of 2 ml and overlayed with one fraction of 30% (w/v) Optiprep™ (9ml) followed by a fraction of 5% (w/v) Optiprep™ (3 ml). Gradients werecentrifuged at 250,000×g for 3 hours at 4° C. and the first 4 ml on top,containing the particles, were collected. After 3 washes with PBS(250,000×g for 2 hours at 4C), pellets were recovered in PBS and storedat −80° C.

Electron microscopy. To visualize MV_(L) and MP on bacteria, S.pneumoniae T4R was grown in C+Y medium until OD₆₀₀=0.4 at 37° C.Bacteria were harvested by centrifugation for 10 min at 4,000×g, 4° C.and pellets were suspended in 100 μl PBS. Glow discharged carbon coatedgrids (Oxford Instruments, UK) were incubated for 1 min with a drop ofbacterial solution or purified MV_(L)/MP preparation and negativelystained with 2% uranyl acetate in water (7 times for 10 sec). Specimenswere examined on a FEI CM120 microscope operated at 80 kV. Images werecollected with a side mounted camera MegaView III (Olympus Soft Imagingsolutions).

Atomic force microscopy. 5 μl of MP samples isolated from S. pneumoniaewere placed onto freshly cleaved mica (Goodfellow Cambridge Ltd.,Cambridge, United Kingdom). The specimens on the mica were blot driedand placed into a desiccator for at least 2 h. Imaging was performed ona Nanoscope IIIa (Digital Instruments, Santa Barbara) Atomic ForceMicroscope using Tapping Mode with standard silicon cantileversoscillating at resonant frequency (270 to 305 kHz). Images werecollected at a scan rate of 0.8-1.5 Hz, depending on sample number andthe size of the scan. The final images were fitted in both axes andpresented in a surface plot of the height mode.

SDS-PAGE and western blotting. The total amount of proteins in purifiedMV_(L) or MP, lysates of A549 cells or dendritic cells stimulated withMV_(L) or MP were quantified with Pierce™ BCA Protein Assay Kit (LifeTechnologies). Samples containing equal amounts of total protein wereresolved by SDS-PAGE using 4-12% Bis-Tris gels (Life Technologies) andtransferred to PVDF membranes. Membranes were then blocked with 5% skimmilk in PBS containing 0.1% Tween-20 and incubated with antibodies asindicated. For detection of pneumolysin a mouse monoclonal antibody(Abcam, final dilution 1:500) was used. Polyclonal GAPDH (1:2,000) andLytA (1:2,000) (3) antisera raised in rabbits were used. Rabbitpolyclonal antibodies against PspC (1:1,000), RrgB (1:1,000), PsaA(1:25,000), PhtD (1:25,000) and mouse polyclonal antibodies against SrtA(1:500) were kindly provided by Novartis Vaccines and Diagnostics. Asloading control for A549 cells and dendritic cell lysates, a mousemonoclonal R-actin antibody (Santa Cruz) was used as primary antibody.Anti-mouse IgG or anti-rabbit IgG conjugated to horseradish peroxidase(GE Healhcare) were used as secondary antibodies (final dilution1:10,000). Blots were developed with Amersham™ ECL Plus Western blottingdetection system (GE Healthcare Life Sciences), using a ChemiDoc™ XRS+(Bio-Rad Laboratories).

Tandem mass spectrometry. Proteins in MV_(L) or MP samples were reduced,alkylated and in-solution digested by trypsin according to a standardoperating procedure. Thereafter the samples were purified by Pierce C18Spin Columns (Thermo Scientific), dried and resolved in 0.1% formicacid. The resulting peptides were separated in reversed-phase on aC18-column and electrosprayed on-line to a Q Exactive Plus massspectrometer (Thermo Finnigan). Tandem mass spectrometry was performedapplying HCD.

Database searches were made using the Sequest algorithm towards a FASTAdatabase including proteins from Streptococcus pneumoniae TIGR4. Thesearch criteria for protein identification were set to at least twomatching peptides of 95% confidence level per protein. Only proteinswith a Sequest score above 20 were considered for analysis, to avoid thepossibility of false positives.

Subcellular localizations of proteins were predicted using the algorithmof website Locate P located at the following URL:www.cmbi.ru.nl/locatep-db/cgi-bin/locatepdb.py (11).

Hemolysis assay. Purified MV_(L) or MP were incubated in 96-well plateswith blood from buffy coats (diluted 1:50 in PBS containing 1 mMdithiothreitol (DTT)) for 1 hour at 37° C. After 50 minutes 1% TritonX-100 in PBS was added to the positive control wells in order to lyseall the erythrocytes. Plates were then spun at 400×g for 15 minutes at4° C.; supernatant was transferred to an optical plate and the opticaldensity was measured at 540 nm.

A549 cell toxicity assay. A459 lung epithelial cells were grown andmaintained at 37° C., with 5% CO₂ in RPMI medium (Gibco) supplementedwith 10% (v/v) Fetal bovine serum (FBS) (HyClone). To assess MV_(L) orMP associated cytotoxic effects, 0.6×10⁶ A549 cells were seeded in6-well plates and incubated overnight at 37° C. Cells were then washedwith PBS and incubated for 24 hours with medium containing MV_(L) or MPat indicated concentrations. Washed cells were labelled with FixableViability Dye eFluor® 780 (1:50,000, eBioscience) for 30 minutes at 4°C. in the dark, in presence or absence of 0.02% NP40 (Sigma) as positivecontrol, and fixed with 4% paraformaldehyde (PFA) for 30 minutes at roomtemperature. Next, cells were gently scraped into PBS containing 1% FBSand analyzed in a Gallios™ Flow Cytometer (Beckman Coulter).

A549 cell uptake assay. To assess MV_(L) and MP uptake by A549 cells viaimmunoblotting, 0.6×10⁶ A549 cells were seeded in 6-well plates andincubated overnight at 37° C. Cells were then washed with PBS andincubated for 24 hours with medium containing the particles at indicatedconcentrations. After washing, cells were lysed in RIPA buffercontaining 1× protease inhibitor cocktail (Roche). Cell lysates wereanalyzed by SDS-PAGE and western blotting.

Immunofluorescence microscopy. A549 cells (6.25×10⁴) were seeded in24-well plates with coverslips on the bottom of the wells, and incubatedat 37° C. overnight. Cells were then washed, fixed with 4% PFA for 30minutes and permeabilized with 1% Triton X-100 in PBS for 5 minutes.MV_(L) or MP were detected with mouse monoclonal anti-Ply (1:200) andpolyclonal rabbit antibodies to LytA (1:200). Respective antigens werevisualized with Alexa Fluor® 488-conjugated goat-anti mouse IgG antibody(Life Technologies) and Alexa Fluor® 350-conjugated goat-anti rabbit IgG(Life Technologies) (1:1,000).

Actin cytoskeleton was stained with Alexa Fluor® 594 Phalloidin (LifeTechnologies, 1:40 dilution) for 1 hour and coverslips were mounted withVectashield (Vector Laboratories, Inc.) on microscope slides. Imageswere acquired with a DeltaVision microscope equipped with a60x-objective. Quick projection images of approximately 20 z-stacks weretaken. Orthogonal views were used to visualize the cell monolayer fromthe x, y, and z axes.

After growth, pneumococci (serotypes 1, 3 and 6B) have been stainedusing the sera from the immunized mice as primary antibody (dilution1:100 in PBS 1% BSA) and Alexa Fluor 488 goat anti mouse (dilution 1:500in PBS 1% BSA) as secondary antibody.

Imaging has been performed with high-resolution Delta Vision EliteSystem using 100x objective (1000×total magnification). FITC Laserintensity 50%, exposure time 0,025 ms (FIGS. 13-15 ).

Quantification of the Signal Detected on the Bacteria afterImmunofluorescence Staining

Using the functions Image>Adjust>Threshold and Analyze>Measure ofImageJ, the area covered by the bacteria and the area covered by thesignal detected on the bacteria after staining with sera were selected,defined and measured. The final signal ratio was calculated by dividingthe area of the signal (detected using the sera) by the total area ofthe bacteria.

Isolation and differentiation of human monocyte-derived dendritic cells.Dendritic cells were isolated using RosetteSep™ Human MonocyteEnrichment Cocktail (Stemcell Technologies) according to themanufacturer's instructions. In brief, blood from buffy coats fromhealthy donors was incubated for 20 minutes with RosetteSep™ HumanMonocyte Enrichment Cocktail (Stemcell Technologies), layered on top ofFicoll-Paque™ Plus (GE Healthcare) and centrifuged at 1200×g for 20minutes without acceleration or brake. The monocyte containing layer wasrecovered, cells washed 7 times with PBS and passed through a 100 μmcell strainer. Monocytes were then differentiated for 6 days in RPMIcontaining 10% FBS supplemented with 37.5 ng/ml of Granulocytemacrophage colony-stimulating factor (GM-CSF) (Peprotech) and 37.5 ng/mlof Interleukin 4 (IL-4) (Peprotech), changing the medium after 4 days.For experiments, cells were resuspended in RPMI containing 10% FBS.

Toxicity and apoptosis assay of dendritic cells. 6×10⁵ cells were seededin 96-well plates and incubated with RPMI containing 10% FBS and MV_(L)or MP at indicated concentrations, or with T4R in a multiplicity ofinfection (MOI) of 20, as positive control, for 24 hours. Gentamicin(100 μg/ml, Sigma) was added after 1 hour of incubation to stop bacteriafrom growing in samples stimulated with T4R. Before staining, cells werewashed once in PBS and once in Annexin V Binding Buffer (BD Pharmingen).Staining was performed with Fixable Viability Dye eFluor® 780 (1:50,000,eBioscience) and FITC Annexin V (1:20, BD Pharmingen) for 30 minutes at4° C., followed by two washes with Annexin buffer. Labelled cells werefixed in 4% PFA for 30 minutes, resuspended in PBS containing 1% FBS andanalyzed in a Gallios™ Flow Cytometer (Beckman Coulter).

Activation assay of dendritic cells. 6×10⁵ cells were seeded in 96-wellplates and incubated in RPMI containing 10% FBS with MV_(L) or MP atindicated concentrations, or with 1 μg/ml lipopolysaccharide (LPS)(Sigma) as positive control, for 24 hours. Cells were stained withPhycoerythrin (PE) Mouse Anti-Human CD86 (BD Pharmingen) and PE-Cy™5Mouse Anti-Human HLA-DR (BD Pharmingen) for 20 minutes at 4° C., washedtwice with PBS and resuspended in PBS 1% FBS. Labelled cells wereanalyzed in a Gallios™ Flow Cytometer (Beckman Coulter).

Uptake assay with dendritic cells. 10⁶ cells were seeded in 96-wellplates and incubated with RPMI containing 10% FBS, in presence orabsence of the inhibitors cytochalasin D (0.5 μg/ml, Sigma) andwortmannin (0.5 μg/ml, Sigma) (C/W), or methyl-R-cyclodextrin (MSCD) (10μM, Sigma), for 30 minutes. Cells were then incubated with particles atindicated concentration for 1 hour and lysed in RIPA buffer containing1× protease inhibitor cocktail (Roche). Cell lysates were analyzed bySDS-PAGE and western blotting.

Quantification of cytokines. Different cytokines were assessed (IL-6,IL-8, IL-10 and TNF) in cell-free supernatants of 10⁵ dendritic cells byEnzyme-Linked Immunosorbent Assay (ELISA), using commercially availableBD OptEIA™ kits from BD Biosciences. Cells were incubated with particlesat indicated concentrations, or with T4R or LPS (as previouslydescribed), for 24 hours.

Opsonophagocytosis assay with RAW cells. RAW 264.7 murine macrophageswere grown and maintained at 37° C., with 5% CO₂ in RPMI medium (Gibco)supplemented with 10% (v/v) Fetal bovine serum (FBS) (HyClone).To assessthe opsonophagocytosis activity of antibodies in immunized mice sera2×10⁵ RAW 264.7 cells were seeded in 24-well plates and incubatedovernight at 37° C. Serotype 1 bacteria were incubated for 30 minutes at37° C. with 5% CO₂ with 20% serum from mice immunized with MP+adjuvant,or with adjuvant alone as negative control. RAW cells were then washedwith PBS and incubated for 1.5 hours with 2.5×10⁷/well of pre-treatedbacteria. Cells were washed three times with PBS to remove unattachedbacteria. To measure total uptake of bacteria, cells were incubated witha 50/50 solution of 2% saponin (Sigma) and trypsin-EDTA (Gibco) for 15minutes at 37° C., to lyse eukaryotic cells, and total bacteria wereplated for enumeration. To evaluate phagocytosis 300 μg/ml of Gentamicin(Sigma) and 0.12 mg/ml of Penicillin G (Sigma) were added to separatewells and incubated 15 minutes at 37° C., to kill extracellularbacteria. Cells were then washed three times with PBS and incubated witha 50/50 solution of 2% saponin and trypsin-EDTA for 15 minutes at 37° C.to lyse eukaryotic cells. To evaluate killing of bacteria insidemacrophages separate wells were treated with antibiotics (as forphagocytosis), washed three times with PBS and then incubated for 1 hourat 37° C. with medium. Cells were then washed three times with PBS andincubated with a 50/50 solution of 2% saponin and trypsin-EDTA for 15minutes at 37° C. to lyse eukaryotic cells.

Mouse IgG ELISA assay. To detect MV- and MP-specific mouse IgG in seraof immunized mice optical plates (Sarstedt) were coated with 1 μg/ml ofMP in 0.1 M Sodium Carbonate buffer pH 9.5, overnight at 4° C. Wellswere then washed three times with PBS containing 0.05% Tween-20 andincubated with PBS with 10% FBS, for 1 hours at room temperature. Afterthree washes wells were incubated with mice sera diluted 1:500 in PBSwith 10% FBS, for 2 hours at room temperature. Wells were then washedthree times and incubated with anti-mouse IgG-HRP (GE Healthcare)diluted 1:500 in PBS 10% FBS, for 2 hours at room temperature. Afterthree washes wells were then incubated with TMB substrate (BDBioscience) for 10 minutes and the reaction was stopped with 1 M H₃PO₄.Absorbance at 450 nm was then measured with a plate reader.

To detect pneumo-specific mouse IgG in sera of immunized mice, bacteriawere grown on blood agar plates overnight at 37° C., resuspended in PBSand heat-inactivated for 2 hours at 60° C. After diluting the bacteriato OD₆₀₀ 0.6, optical plates (Sarstedt) were coated with 100 μl ofbacteria in 0.1 M Sodium Carbonate buffer pH 9.5, overnight at 4° C.Wells were then washed three times with PBS and incubated with PBS with2.5% skim milk, for 2 hours at room temperature. After three washeswells were incubated with mice sera diluted 1:500 in PBS, for 1 hour atroom temperature. Wells were then washed three times and incubated withanti-mouse IgG-HRP (GE Healthcare) diluted 1:500 in PBS, for 1 hour atroom temperature. After three washes well were then incubated with TMBsubstrate (BD Bioscience) for 10 minutes and the reaction was stoppedwith 1 M H₃PO₄. Absorbance at 450 nm was then measured with a platereader.

Statistical analysis. For multiple comparisons the nonparametric ANOVAtest was used to assess the presence of the differences between thegroups, then the Dunn's test was used to make pairwise comparisons; fortwo groups comparison the non-parametric two tailed Wilcoxon's rank sumtest (also known as Mann-Whitney test) was used. Statisticallysignificant data was defined as *P<0.05, **P<0.01, ***P<0.001,****=P<0.0001.

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The invention claimed is:
 1. A method for inducing protective immunityagainst Streptococcus pneumoniae in a subject, comprising administeringto the subject a composition comprising a Streptococcus pneumoniaemembrane vesicle microparticle (MP), wherein said MP comprises: i. theprotein Ply at the level of ≥0.070 μg/μg total protein in the MP; ii.the protein LytA at the level of ≥0.070 μg/μg total protein in the MP;iii. the protein PspC at the level of ≥0.130 μg/μg total protein in theMP; or iv. the protein RrgB at the level of ≥0.020 μg/μg total proteinin the MP.
 2. The method according to claim 1, wherein the immunity isprotective against a condition selected from pneumococcal sinusitis,pneumococcal otitis, pneumococcal pneumonia, and invasive pneumococcaldisease including but not limited to pneumococcal sepsis andpneumococcal meningitis.
 3. The method according to claim 1, wherein thecomposition is a liquid and comprises MPs in an amount of at least 1μg/ml.
 4. The method according to claim 1, wherein said MP comprises: i.the protein Ply at the level of ≥0.070 μg/μg total protein in the MP;ii. the protein LytA at the level of ≥0.070 μg/μg total protein in theMP; iii. the protein PspC at the level of ≥0.130 μg/μg total protein inthe MP; and iv. the protein RrgB at the level of ≥0.020 μg/μg totalprotein in the MP.
 5. The method according to claim 1, wherein the MP is5-300 nm in diameter.
 6. The method according to claim 1, wherein the MPis 10-125 nm in diameter.
 7. The method according to claim 1, whereinthe composition elicits antibodies against Streptococcus pneumoniaeserotype 3 when administered to a mammalian host.
 8. The methodaccording to claim 1, wherein the composition elicits serotypeindependent antibodies against Streptococcus pneumoniae whenadministered to a mammalian host.
 9. The method according to claim 1,wherein the composition is administered intranasally.
 10. The methodaccording to claim 1, wherein the composition further comprises anadjuvant.
 11. The method according to claim 10, wherein the adjuvantcomprises aluminium hydroxide.
 12. The method according to claim 2,wherein the immunity is protective against invasive pneumococcaldisease.
 13. The method according to claim 1, wherein the MP comprises acapsular polysaccharide of a capsular serotype of Streptococcuspneumoniae at a level of ≥0.001 μg/μg total protein in the MP.
 14. Themethod according to claim 1, wherein said MP comprises the protein Plyat the level of ≥0.070 μg/μg total protein in the MP.
 15. The methodaccording to claim 1, wherein said MP comprises the protein LytA at thelevel of ≥0.070 μg/μg total protein in the MP.
 16. The method accordingto claim 1, wherein said MP comprises the protein PspC at the level of≥0.130 μg/μg total protein in the MP.
 17. The method according to claim1, wherein said MP comprises the protein RrgB at the level of ≥0.020μg/μg total protein in the MP.
 18. The method according to claim 1,wherein said MP comprises the protein Ply at the level of ≥0.35 μg/μgtotal protein in the MP.
 19. The method according to claim 1, whereinsaid MP comprises the protein LytA at the level of ≥0.20 μg/μg totalprotein in the MP.
 20. The method according to claim 1, wherein said MPcomprises the protein PspC at the level of ≥0.3 μg/μg total protein inthe MP.
 21. The method according to claim 1, wherein said MP comprisesthe protein RrgB at the level of ≥0.028 μg/μg total protein in the MP.